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Michael Philips

Michael Philips

Gene Regulation in Eukaryotes

Slide Duration:

Table of Contents

I. The Beginnings of Molecular Biology
Biochemistry Review: Importance of Chemical Bonds

53m 29s

Intro
0:00
Lesson Overview
0:14
Chemical Bonds
0:41
Attractive Forces That Hold Atoms Together
0:44
Types of Bonds
0:56
Covalent Bonds
1:34
Valence Number
1:58
H O N C P S Example
2:50
Polar Bonds
7:23
Non-Polar Bond
8:46
Non-Covalent Bonds
9:46
Ionic Bonds
10:25
Hydrogen Bonds
10:52
Hydrophobic Interactions
11:34
Van Der Waals Forces
11:58
Example 1
12:51
Properties of Water
18:27
Polar Molecule
13:34
H-bonding Between Water H20 Molecules
19:29
Hydrophobic Interactions
20:30
Chemical Reactions and Free Energy
22:52
Transition State
23:00
What Affect the Rate
23:27
Forward and Reserve Reactions Occur Simultaneously But at Different Rate
23:51
Equilibrium State
24:29
Equilibrium Constant
25:18
Example 2
26:16
Chemical Reactions and Free Energy
27:49
Activation Energy
28:00
Energy Barrier
28:22
Enzymes Accelerate Reactions by Decreasing the Activation Energy
29:04
Enzymes Do Not Affect the Reaction Equilibrium or the Change in Free Energy
29:22
Gibbs Free Energy Change
30:50
Spontaneity
31:18
Gibbs Free Energy Change Determines Final Concentrations of Reactants
34:36
Endodermic vs. Exothermic Graph
35:00
Example 3
38:46
Properties of DNA
39:37
Antiparallel Orientation
40:29
Purine Bases Always Pairs Pyrimidine Bases
41:15
Structure Images
42:36
A, B, Z Forms
43:33
Major and Minor Grooves
44:09
Hydrogen Bonding and Hydrophobic Interactions Hold the Two Strands Together
44:39
Denaturation and Renaturation of DNA
44:56
Ways to Denature dsDNA
45:28
Renature When Environment is Brought Back to Normal
46:05
Hyperchromiicity
46:36
Absorbs UV Light
47:01
Spectrophotometer
48:01
Graph Example?
49:05
Example 4
51:02
Mendelian Genetics & Foundational Experiments

1h 9m 27s

Intro
0:00
Lesson Overview
0:22
Gregor Johann Mendel
1:01
Was a Biologist and Botanist
1:14
Published Seminal Paper on Hybridization and Inheritance in the Pea Plant
1:20
Results Criticized
1:28
Father of Modern Genetics
1:59
Mendel’s Laws
2:19
1st Law: Principle of Independent Segregation of Alleles
2:27
2nd Law: Principle of Independent Assortment of Genes
2:34
Principle of Independent Segregation (of Alleles)
2:41
True Breeding Lines / Homozygous
2:42
Individuals Phenotypes Determined by Genes
3:15
Alleles
3:37
Alleles Can Be Dominant or Recessive
3:50
Genotypes Can be Experimentally Determined by Mating and Analyzing the Progeny
5:36
Individual Alleles Segregate Independently Into Gametes
5:55
Example 1
6:18
Principle of Independent Segregation (of Alleles)
16:11
Individual Genes Sort Independently Into Gametes
16:22
Each Gamete Receives One Allele of Each Gene: 50/50 Chance
16:46
Genes Act Independently to Determine Unrelated Phenotypes
16:57
Example: Punnett Square
17:15
Example 2
21:36
The Chromosomal Theory of Inheritance
30:41
Walter S Sutton Linked Cytological Studies with Mendels Work
31:02
Diploid Cells Have Two Morphologically Similar Sets of Chromosomes and Each Haploid Gamete Receives One Set
31:17
Genes Are on Chromosome
31:33
Gene for Seed Color’s on a Different Chromosome Than Gene for Seed Texture
31:44
Gene Linkage
31:55
Mendel’s 2nd Law
31:57
Genes Said to Be Linked To Each Other
32:09
Linkage Between Genes
32:29
Linkage is Never 100% Complete
32:41
Genes are Found on Chromosomes
33:00
Thomas Hunt Morgan and Drosophila Melanogaster
33:01
Mutation Linked to X Chromosome
33:15
Linkage of White Gene
33:23
Eye Color of Progeny Depended on Sex of Parent
33:34
Y Chromosome Does Not Carry Copy of White Gene
33:44
X Linked Genes, Allele is Expressed in Males
33:56
Example
34:11
Example 3
35:52
Discovery of the Genetic Material of the Cell
41:52
Transforming Principle
42:44
Experiment with Streptococcus Pneumoniae
42:55
Beadle and Tatum Proposed Genes Direct the Synthesis of Enzymes
45:15
One Gene One Enzyme Hypothesis
45:46
One Gene One Polypeptide Theory
45:52
Showing the Transforming Material was DNA
46:14
Did This by Fractionating Heat-Killed “S” Strains into DNA, RNA, and Protein
46:32
Result: Only the DNA Fraction Could Transform
47:15
Leven: Tetranucleotide Hypothesis
48:00
Chargaff Showed This Was Not the Case
48:48
Chargaff: DNA of Different Species Have Different Nucleotide Composition
49:02
Hershey and Chase: DNA is the Genetic Material
50:02
Incorporate Sulfur into Protein and Phosphorous into DNA
51:12
Results: Phosphorase Entered Bacteria and Progeny Phage, But no Sulfur
53:11
Rosalind Franklin’s “Photo 51” Showing the Diffraction Pattern of DNA
53:50
Watson and Crick: Double Helical Structure of DNA
54:57
Example 4
56:56
Discovery of the Genetic Material of the Cell
58:09
Kornberg: DNA Polymerase I
58:10
Three Postulated Methods of DNA Replication
59:22
Meselson and Stahl: DNA Replication is Semi-Conservative
1:00:21
How DNA Was Made Denser
1:00:52
Discovery of RNA
1:03:32
Ribosomal RNA
1:03:48
Transfer RNA
1:04:00
Messenger RNA
1:04:30
The Central Dogma of Molecular Biology
1:04:49
DNA and Replication
1:05:08
DNA and Transcription = RNA
1:05:26
RNA and Translation = Protein
1:05:41
Reverse Transcription
1:06:08
Cracking the Genetic Code
1:06:58
What is the Genetic Code?
1:07:04
Nirenberg Discovered the First DNA Triplet That Would Make an Amino Acid
1:07:16
Code Finished in 1966 and There Are 64 Possibilities or Triplet Repeats/ Codons
1:07:54
Degeneracy of the Code
1:08:53
II. Structure of Macromolecules
Structure of Proteins

49m 44s

Intro
0:00
Lesson Overview
0:10
Amino Acids
0:47
Structure
0:55
Acid Association Constant
1:55
Amino Acids Make Up Proteins
2:15
Table of 21 Amino Acid Found in Proteins
3:34
Ionization
5:55
Cation
6:08
Zwitterion
7:51
Anion
9:15
Example 1
10:53
Amino Acids
13:11
L Alpha Amino Acids
13:19
Only L Amino Acids Become Incorporated into Proteins
13:28
Example 2
13:46
Amino Acids
18:20
Non-Polar
18:41
Polar
18:58
Hydroxyl
19:52
Sulfhydryl
20:21
Glycoproteins
20:41
Pyrrolidine
21:30
Peptide (Amide) Bonds
22:18
Levels of Organization
23:35
Primary Structure
23:54
Secondary Structure
24:22
Tertiary Structure
24:58
Quaternary Structure
25:27
Primary Structure: Specific Amino Acid Sequence
25:54
Example 3
27:30
Levels of Organization
29:31
Secondary Structure: Local 3D
29:32
Example 4
30:37
Levels of Organization
32:59
Tertiary Structure: Total 3D Structure of Protein
33:00
Quaternary Structure: More Than One Subunit
34:14
Example 5
34:52
Protein Folding
37:04
Post-Translational Modifications
38:21
Can Alter a Protein After It Leaves the Ribosome
38:33
Regulate Activity, Localization and Interaction with Other Molecules
38:52
Common Types of PTM
39:08
Protein Classification
40:22
Ligand Binding, Enzyme, DNA or RNA Binding
40:36
All Other Functions
40:53
Some Functions: Contraction, Transport, Hormones, Storage
41:34
Enzymes as Biological Catalysts
41:58
Most Metabolic Processes Require Catalysts
42:00
Most Biological Catalysts Are Proteins
43:13
Enzymes Have Specificity of Reactants
43:33
Enzymes Have an Optimum pH and Temperature
44:31
Example 6
45:08
Structure of Nucleic Acids

1h 2m 10s

Intro
0:00
Lesson Overview
0:06
Nucleic Acids
0:26
Biopolymers Essential for All Known Forms of Life That Are Composed of Nucleotides
0:27
Nucleotides Are Composed of These
1:17
Nucleic Acids Are Bound Inside Cells
2:10
Nitrogen Bases
2:49
Purines
3:01
Adenine
3:10
Guanine
3:20
Pyrimidines
3:54
Cytosine
4:25
Thymine
4:33
Uracil
4:42
Pentoses
6:23
Ribose
6:45
2' Deoxyribose
6:59
Nucleotides
8:43
Nucleoside
8:56
Nucleotide
9:16
Example 1
10:23
Polynucleotide Chains
12:18
What RNA and DNA Are Composed of
12:37
Hydrogen Bonding in DNA Structure
13:55
Ribose and 2! Deoxyribose
14:14
DNA Grooves
14:28
Major Groove
14:46
Minor Groove
15:00
Example 2
15:20
Properties of DNA
24:15
Antiparallel Orientation
24:25
Phosphodiester Linkage
24:50
Phosphate and Hydroxyl Group
25:05
Purine Bases Always Pairs Pyramidine Bases
25:30
A, B, Z Forms
25:55
Major and Minor Grooves
26:24
Hydrogen Bonding and Hydrophobic Interactions Hold Strands Together
26:34
DNA Topology - Linking Number
27:14
Linking Number
27:31
Twist
27:57
Writhe
28:31
DNA Topology - Supercoiling
31:50
Example 3
33:16
III. Maintenance of the Genome
Genome Organization: Chromatin & Nucleosomes

57m 2s

Intro
0:00
Lesson Overview
0:09
Quick Glossary
0:24
DNA
0:29
Gene
0:34
Nucleosome
0:47
Chromatin
1:07
Chromosome
1:19
Genome
1:30
Genome Organization
1:38
Physically Cellular Differences
3:09
Eukaryotes
3:18
Prokaryotes, Viruses, Proteins, Small Molecules, Atoms
4:06
Genome Variance
4:27
Humans
4:52
Junk DNA
5:10
Genes Compose Less Than 40% of DNA
6:03
Chart
6:26
Example 1
8:32
Chromosome Variance - Size, Number, and Density
10:27
Chromosome
10:47
Graph of Human Chromosomes
10:58
Eukaryotic Cell Cycle
12:07
Requirements for Proper Chromosome Duplication and Segregation
13:07
Centromeres and Telomeres
13:28
Origins of Replication
13:38
Illustration: Chromosome
13:44
Chromosome Condensation
15:52
Naked DNA to Start
16:00
Beads on a String
16:13
Mitosis
16:52
Start with Two Different Chromosomes
17:18
Split Into Two Diploid Cells
17:26
Prophase
17:42
Prometaphase
17:52
Metaphase
19:10
Anaphase
19:27
Telophase
20:11
Cytokinesis
20:31
Cohesin and Condensis
21:06
Illustration: Cohesin and Condensis
21:19
Cohesin
21:38
Condensin
21:43
Illustration of What Happens
21:50
Cohesins
27:23
Loaded During Replication and Cleaved During Mitosis
27:30
Separase
27:36
Nucleosomes
27:59
Histone Core
28:50
Eight Histone Proteins
28:57
Octamer of Core Histones Picture
29:14
Chromosome Condensation via H1
30:59
Allows Transition to Compact DNA
31:09
When Not in Mitosis
31:37
Histones Decrease Available Binding Sites
32:38
Histone Tails
33:21
Histone Code
35:32
Epigenetic Code
35:56
Phosphorylation
36:45
Acetylation
36:57
Methylation
37:01
Ubiquitnation
37:04
Example 2
38:48
Nucleosome Assembly
41:22
Duplication of DNA Requires Duplication of Histones
41:50
Old Histones Are Recycled
42:00
Parental H3-H4 Tetramers Facilitate the Inheritance of Chromatin States
44:04
Example 3
46:00
Chromatin Remodeling
48:12
Example 4
53:28
DNA Replication

1h 9m 55s

Intro
0:00
Lesson Overview
0:06
Eukaryotic Cell Cycle
0:50
G1 Growth Phase
0:57
S Phase: DNA & Replication
1:09
G2 Growth Phase
1:28
Mitosis
1:36
Normal Human Cell Divides About Every 24 Hours
1:40
Eukaryotic DNA Replication
2:04
Watson and Crick
2:05
Specific Base Pairing
2:37
DNA Looked Like Tetrinucleotide
2:55
What DNA Looks Like Now
3:18
Eukaryotic DNA Replication - Initiation
3:44
Initiation of Replication
3:53
Primer Template Junction
4:25
Origin Recognition Complex
7:00
Complex of Proteins That Recognize the Proper DNA Sequence for Initiation of Replication
7:35
Prokaryotic Replication
7:56
Illustration
8:54
DNA Helicases (MCM 2-7)
11:53
Eukaryotic DNA Replication
14:36
Single-Stranded DNA Binding Proteins
14:59
Supercoils
16:30
Topoisomerases
17:35
Illustration with Helicase
19:05
Synthesis of the RNA Primer by DNA Polymerase Alpha
20:21
Subunit: Primase RNA Polymerase That Synthesizes the RNA Primer De Navo
20:38
Polymerase Alpha-DNA Polymerase
21:01
Illustration of Primase Function Catalyzed by DnaG in Prokaryotes
21:22
Recap
24:02
Eukaryotic DNA Replication - Leading Strand
25:02
Synthesized by DNA Polymerase Epsilon
25:08
Proof Reading
25:26
Processivity Increased by Association with PCNA
25:47
What is Processivity?
26:19
Illustration: Write It Out
27:03
The Lagging Strand/ Discontinuing Strand
30:52
Example 1
31:57
Eukaryotic DNA Replication - Lagging Strand
32:46
Discontinuous
32:55
DNA Polymerase Delta
33:15
Okazaki Fragments
33:36
Illustration
33:55
Eukaryotic DNA Replication - Okazaki Fragment Processing
38:26
Illustration
38:44
When Does Okazaki Fragments Happen
40:32
Okazaki Fragments Processing
40:41
Illustration with Okazaki Fragments Process Happening
41:13
Example 2
47:42
Example 3
49:20
Telomeres
56:01
Region of Repetitive Nucleotide Sequences
56:26
Telomeres Act as Chromosome Caps by Binding Proteins
57:42
Telomeres and the End Replication Problem
59:56
Need to Use a Primer
59:57
DNA Mutations & Repairs

1h 13m 8s

Intro
0:00
Lesson Overview
0:06
Damage vs. Mutation
0:40
DNA Damage-Alteration of the Chemical Structure of DNA
0:45
DNA Mutation-Permanent Change of the Nucleotide Sequence
1:01
Insertions or Deletions (INDELS)
1:22
Classes of DNA Mutations
1:50
Spontaneous Mutations
2:00
Induced Mutations
2:33
Spontaneous Mutations
3:21
Tautomerism
3:28
Depurination
4:09
Deamination
4:30
Slippage
5:44
Induced Mutations - Causes
6:17
Chemicals
6:24
Radiation
7:46
Example 1
8:30
DNA Mutations - Tobacco Smoke
9:59
Covalent Adduct Between DNA and Benzopyrene
10:02
Benzopyrene
10:20
DNA Mutations - UV Damage
12:16
Oxidative Damage from UVA
12:30
Thymidine Dimer
12:34
Example 2
13:33
DNA Mutations - Diseases
17:25
DNA Repair
18:28
Mismatch Repair
19:15
How to Recognize Which is the Error: Recognize Parental Strand
22:23
Example 3
26:54
DNA Repair
32:45
Damage Reversal
32:46
Base-Excision Repair (BER)
34:31
Example 4
36:09
DNA Repair
45:43
Nucleotide Excision Repair (NER)
45:48
Nucleotide Excision Repair (NER) - E.coli
47:51
Nucleotide Excision Repair (NER) - Eukaryotes
50:29
Global Genome NER
50:47
Transcription Coupled NER
51:01
Comparing MMR and NER
51:58
Translesion Synthesis (TLS)
54:40
Not Really a DNA Repair Process, More of a Damage Tolerance Mechanism
54:50
Allows Replication Past DNA Lesions by Polymerase Switching
55:20
Uses Low Fidelity Polymerases
56:27
Steps of TLS
57:47
DNA Repair
1:00:37
Recombinational Repair
1:00:54
Caused By Ionizing Radiation
1:00:59
Repaired By Three Mechanisms
1:01:16
Form Rarely But Catastrophic If Not Repaired
1:01:42
Non-homologous End Joining Does Not Require Homology To Repair the DSB
1:03:42
Alternative End Joining
1:05:07
Homologous Recombination
1:07:41
Example 5
1:09:37
Homologous Recombination & Site-Specific Recombination of DNA

1h 14m 27s

Intro
0:00
Lesson Overview
0:16
Homologous Recombination
0:49
Genetic Recombination in Which Nucleotide Sequences Are Exchanged Between Two Similar or Identical Molecules of DNA
0:57
Produces New Combinations of DNA Sequences During Meiosis
1:13
Used in Horizontal Gene Transfer
1:19
Non-Crossover Products
1:48
Repairs Double Strand Breaks During S/Gs
2:08
MRN Complex Binds to DNA
3:17
Prime Resection
3:30
Other Proteins Bind
3:40
Homology Searching and subsequent Strand Invasion by the Filament into DNA Duplex
3:59
Holliday Junction
4:47
DSBR and SDSA
5:44
Double-Strand Break Repair Pathway- Double Holliday Junction Model
6:02
DSBR Pathway is Unique
6:11
Converted Into Recombination Products by Endonucleases
6:24
Crossover
6:39
Example 1
7:01
Example 2
8:48
Double-Strand Break Repair Pathway- Synthesis Dependent Strand Annealing
32:02
Homologous Recombination via the SDSA Pathway
32:20
Results in Non-Crossover Products
32:26
Holliday Junction is Resolved via Branch Migration
32:43
Example 3
34:01
Homologous Recombination - Single Strand Annealing
42:36
SSA Pathway of HR Repairs Double-Strand Breaks Between Two Repeat Sequences
42:37
Does Not Require a Separate Similar or Identical Molecule of DNA
43:04
Only Requires a Single DNA Duplex
43:25
Considered Mutagenic Since It Results in Large Deletions of DNA
43:42
Coated with RPA Protein
43:58
Rad52 Binds Each of the Repeated Sequences
44:28
Leftover Non-Homologous Flaps Are Cut Away
44:37
New DNA Synthesis Fills in Any Gaps
44:46
DNA Between the Repeats is Always Lost
44:55
Example 4
45:07
Homologous Recombination - Break Induced Replication
51:25
BIR Pathway Repairs DSBs Encountered at Replication Forks
51:34
Exact Mechanisms of the BIR Pathway Remain Unclear
51:49
The BIR Pathway Can Also Help to Maintain the Length of Telomeres
52:09
Meiotic Recombination
52:24
Homologous Recombination is Required for Proper Chromosome Alignment and Segregation
52:25
Double HJs are Always Resolved as Crossovers
52:42
Illustration
52:51
Spo11 Makes a Targeted DSB at Recombination Hotspots
56:30
Resection by MRN Complex
57:01
Rad51 and Dmc1 Coat ssDNA and Promote Strand Invasion and Holliday Junction Formation
57:04
Holliday Junction Migration Can Result in Heteroduplex DNA Containing One or More Mismatches
57:22
Gene Conversion May Result in Non-Mendelian Segregation
57:36
Double-Strand Break Repair in Prokaryotes - RecBCD Pathway
58:04
RecBCD Binds to and Unwinds a Double Stranded DNA
58:32
Two Tail Results Anneal to Produce a Second ssDNA Loop
58:55
Chi Hotspot Sequence
59:40
Unwind Further to Produce Long 3 Prime with Chi Sequence
59:54
RecBCD Disassemble
1:00:23
RecA Promotes Strand Invasion - Homologous Duplex
1:00:36
Holliday Junction
1:00:50
Comparison of Prokaryotic and Eukaryotic Recombination
1:01:49
Site-Specific Recombination
1:02:41
Conservative Site-Specific Recombination
1:03:10
Transposition
1:03:46
Transposons
1:04:12
Transposases Cleave Both Ends of the Transposon in Original Site and Catalyze Integration Into a Random Target Site
1:04:21
Cut and Paste
1:04:37
Copy and Paste
1:05:36
More Than 40% of Entire Human Genome is Composed of Repeated Sequences
1:06:15
Example 5
1:07:14
IV. Gene Expression
Transcription

1h 19m 28s

Intro
0:00
Lesson Overview
0:07
Eukaryotic Transcription
0:27
Process of Making RNA from DNA
0:33
First Step of Gene Expression
0:50
Three Step Process
1:06
Illustration of Transcription Bubble
1:17
Transcription Starting Site is +1
5:15
Transcription Unit Extends From the Promoter to the Termination Region
5:40
Example 1
6:03
Eukaryotic Transcription: Initiation
14:27
RNA Polymerase II Binds to TATA Box to Initiate RNA Synthesis
14:34
TATA Binding Protein Binds the TATA Box
14:50
TBP Associated Factors Bind
15:01
General Transcription Factors
15:22
Initiation Complex
15:30
Example 2
15:44
Eukaryotic Transcription
17:59
Elongation
18:07
FACT (Protein Dimer)
18:24
Eukaryotic Transcription: Termination
19:36
Polyadenylation is Linked to Termination
19:42
Poly-A Signals Near the End of the pre-mRNA Recruit to Bind and Cleave mRNA
20:00
Mature mRNA
20:27
Dissociate from Template DNA Strand
21:13
Example 3
21:53
Eukaryotic Transcription
25:49
RNA Polymerase I Transcribes a Single Gene That Encodes a Long rRNA Precursor
26:14
RNA Polymerase III Synthesizes tRNA, 5S rRNA, and Other Small ncRNA
29:11
Prokaryotic Transcription
32:04
Only One Multi-Subunit RNA Polymerase
32:38
Transcription and Translation Occurs Simultaneously
33:41
Prokaryotic Transcription - Initiation
38:18
Initial Binding Site
38:33
Pribnox Box
38:42
Prokaryotic Transcription - Elongation
39:15
Unwind Helix and Expand Replication Bubble
39:19
Synthesizes DNA
39:35
Sigma 70 Subunit is Released
39:50
Elongation Continues Until a Termination Sequence is Reached
40:08
Termination - Prokaryotes
40:17
Example 4
40:30
Example 5
43:58
Post-Transcriptional Modifications
47:15
Can Post Transcribe your rRNA, tRNA, mRNA
47:28
One Thing In Common
47:38
RNA Processing
47:51
Ribosomal RNA
47:52
Transfer RNA
49:08
Messenger RNA
50:41
RNA Processing - Capping
52:09
When Does Capping Occur
52:20
First RNA Processing Event
52:30
RNA Processing - Splicing
53:00
Process of Removing Introns and Rejoining Exons
53:01
Form Small Nuclear Ribonucleoproteins
53:46
Example 6
57:48
Alternative Splicing
1:00:06
Regulatory Gene Expression Process
1:00:27
Example
1:00:42
Example 7
1:02:53
Example 8
1:09:36
RNA Editing
1:11:06
Guide RNAs
1:11:25
Deamination
1:11:52
Example 9
1:13:50
Translation

1h 15m 1s

Intro
0:00
Lesson Overview
0:06
Linking Transcription to Translation
0:39
Making RNA from DNA
0:40
Occurs in Nucleus
0:59
Process of Synthesizing a Polypeptide from an mRNA Transcript
1:09
Codon
1:43
Overview of Translation
4:54
Ribosome Binding to an mRNA Searching for a START Codon
5:02
Charged tRNAs will Base Pair to mRNA via the Anticodon and Codon
5:37
Amino Acids Transferred and Linked to Peptide Bond
6:08
Spent tRNAs are Released
6:31
Process Continues Until a STOP Codon is Reached
6:55
Ribosome and Ribosomal Subunits
7:55
What Are Ribosomes?
8:03
Prokaryotes
8:42
Eukaryotes
10:06
Aminoacyl Site, Peptidyl tRNA Site, Empty Site
10:51
Major Steps of Translation
11:35
Charing of tRNA
11:37
Initiation
12:48
Elongation
13:09
Termination
13:47
“Charging” of tRNA
14:35
Aminoacyl-tRNA Synthetase
14:36
Class I
16:40
Class II
16:52
Important About This Reaction: It Is Highly Specific
17:10
ATP Energy is Required
18:42
Translation Initiation - Prokaryotes
18:56
Initiation Factor 3 Binds at the E-Site
19:09
Initiation Factor 1 Binds at the A-Site
20:15
Initiation Factor 2 and GTP Binds IF1
20:50
30S Subunit Associates with mRNA
21:05
N-Formyl-met-tRNA
22:34
Complete 30S Initiation Complex
23:49
IF3 Released and 50S Subunit Binds
24:07
IF1 and IF2 Released Yielding a Complete 70S Initiation Complex
24:24
Deformylase Removes Formyl Group
24:45
Example 1
25:11
Translation Initiation - Eukaryotes
29:35
Small Subunit is Already Associated with the Initiation tRNA
29:47
Formation of 43S Pre-Initiation Complex
30:02
Circularization of mRNA by eIF4
31:05
48S Pre-Initiation Complex
35:47
Example 2
38:57
Translation - Elongation
44:00
Charging, Initiation, Elongation, Termination All Happens Once
44:14
Incoming Charged tRNA Binds the Complementary Codon
44:31
Peptide Bond Formation
45:06
Translocation Occurs
46:05
tRNA Released
46:51
Example 3
47:11
Translation - Termination
55:26
Release Factors Terminate Translation When Ribosomes Come to a Stop Codon
55:38
Release Factors Are Proteins, Not tRNAs, and Do Not Carry an Amino Acid
55:50
Class I Release Factors
55:16
Class II Release Factors
57:03
Example 4
57:40
Review of Translation
1:01:15
Consequences of Altering the Genetic Code
1:02:40
Silent Mutations
1:03:37
Missense Mutations
1:04:24
Nonsense Mutations
1:05:28
Genetic Code
1:06:40
Consequences of Altering the Genetic Code
1:07:43
Frameshift Mutations
1:07:55
Sequence Example
1:08:07
V. Gene Regulation
Gene Regulation in Prokaryotes

45m 40s

Intro
0:00
Lesson Overview
0:08
Gene Regulation
0:50
Transcriptional Regulation
1:01
Regulatory Proteins Control Gene Expression
1:18
Bacterial Operons-Lac
1:58
Operon
2:02
Lactose Operon in E. Coli
2:31
Example 1
3:33
Lac Operon Genes
7:19
LacZ
7:25
LacY
7:40
LacA
7:55
LacI
8:10
Example 2
8:58
Bacterial Operons-Trp
17:47
Purpose is to Produce Trptophan
17:58
Regulated at Initiation Step of Transcription
18:04
Five Genes
18:07
Derepressible
18:11
Example 3
18:32
Bacteriophage Lambda
28:11
Virus That Infects E. Coli
28:24
Temperate Lifecycle
28:33
Example 4
30:34
Regulation of Translation
39:42
Binding of RNA by Proteins Near the Ribosome- Binding Site of the RNA
39:53
Intramolecular Base Pairing of mRNA to Hide Ribosome Binding Site
40:14
Post-transcriptional Regulation of rRNA
40:35
Example 5
40:08
Gene Regulation in Eukaryotes

1h 6m 6s

Intro
0:00
Lesson Overview
0:06
Eukaryotic Transcriptional Regulations
0:18
Transcription Factors
0:25
Insulator Protein
0:55
Example 1
1:44
Locus Control Regions
4:00
Illustration
4:06
Long Range Regulatory Elements That Enhance Expressions of Linked Genes
5:40
Allows Order Transcription of Downstream Genes
6:07
(Ligand) Signal Transduction
8:12
Occurs When an Extracellular Signaling Molecule Activates a Specific Receptor Located on the Cell
8:19
Examples
9:10
N F Kappa B
10:01
Dimeric Protein That Controls Transcription
10:02
Ligands
10:29
Example 2
11:04
JAK/ STAT Pathway
13:19
Turned on by a Cytokine
13:23
What is JAK
13:34
What is STAT
13:58
Illustration
14:38
Example 3
17:00
Seven-Spanner Receptors
20:49
Illustration: What Is It
21:01
Ligand Binding That Is Activating a Process
21:46
How This Happens
22:17
Example 4
24:23
Nuclear Receptor Proteins (NRPs)
28:45
Sense Steroid and Thyroid Hormones
28:56
Steroid Hormones Bind Cytoplasmic NRP Homodimer
29:10
Hormone Binds NRP Heterodimers Already Present in the Nucleus
30:11
Unbound Heterodimeric NRPs Can Cause Deacetylation of Lysines of Histone Tails
30:54
RNA Interference
32:01
RNA Induced Silencing Complex (RISC)
32:39
RNAi
33:54
RISC Pathway
34:34
Activated RISC Complex
34:41
Process
34:55
Example
39:27
Translational Regulation
41:17
Global Regulation
41:37
Competitive Binding of 5 Prime CAP of mRNA
42:34
Translation-Dependent Regulation
44:56
Nonsense Mediated mRNA Decay
45:23
Nonstop Mediated mRNA Decay
46:17
Epigenetics
48:53
Inherited Patterns of Gene Expression Resulting from Chromatin Alteration
49:15
Three Ways to Happen
50:17
DNA Sequence Does Not Act Alone in Passing Genetic Information to Future Generations
50:30
DNA Methylation
50:57
Occurs at CpG Sites Via DNA Methyltransferase Enzymes
50:58
CpG Islands Are Regions with a High Frequency of CpG Sites
52:49
Methylation of Multiple CpG Sites Silence Nearby Gene Transcription
53:32
DNA Methylation
53:46
Pattern Can Be Passed to Daughter Cells
53:47
Prevents SP1 Transcription Factors From Binding to CpG Island
54:02
MECP2
54:10
Example 5
55:27
Nucleosomes
56:48
Histone Core
57:00
Histone Protein
57:03
Chromosome Condensation Via J1
57:32
Linker Histone H1
57:33
Compact DNA
57:37
Histone Code
57:54
Post-translational Modifications of N-Terminal Histone Tails is Part of the Epigenetic Code
57:55
Phosphorylation, Acetylation, Methylation, Ubiquitination
58:09
Example 6
58:52
Nucleosome Assembly
59:13
Duplication of DNA Requires Duplication of Histones by New Protein Synthesis
59:14
Old Histones are Recycled
59:24
Parental H3-H4 Tetramers
58:57
Example 7
1:00:05
Chromatin Remodeling
1:01:48
Example 8
1:02:36
Transcriptionally Repressed State
1:02:45
Acetylation of Histones
1:02:54
Polycomb Repressors
1:03:19
PRC2 Protein Complex
1:03:38
PRC1 Protein Complex
1:04:02
MLL Protein Complex
1:04:09
VI. Biotechnology and Applications to Medicine
Basic Molecular Biology Research Techniques

1h 8m 41s

Intro
0:00
Lesson Overview
0:10
Gel Electraophoresis
0:31
What is Gel Electraophoresis
0:33
Nucleic Acids
0:50
Gel Matrix
1:41
Topology
2:18
Example 1
2:50
Restriction Endonucleases
8:07
Produced by Bacteria
8:08
Sequence Specific DNA Binding Proteins
8:36
Blunt or Overhanging Sticky Ends
9:04
Length Determines Approximate Cleavage Frequency
10:30
Cloning
11:18
What is Cloning
11:29
How It Works
12:12
Ampicillin Example
12:55
Example 2
13:19
Creating a Genomic DNA Library
19:33
Library Prep
19:35
DNA is Cut to Appropriate Sizes and Ligated Into Vector
20:04
Cloning
20:11
Transform Bacteria
20:19
Total Collection Represents the Whole Genome
20:29
Polymerase Chain Reaction
20:54
Molecular Biology Technique to Amplify a Small Number of DNA Molecules to Millions of Copies
21:04
Automated Process Now
21:22
Taq Polymerase and Thermocycler
21:38
Molecular Requirements
22:32
Steps of PCR
23:40
Example 3
24:42
Example 4
34:45
Southern Blot
35:25
Detect DNA
35:44
How It Works
35:50
Western Blot
37:13
Detects Proteins of Interest
37:14
How It Works
37:20
Northern Blot
39:08
Detects an RNA Sequence of Interest
39:09
How It Works
39:21
Illustration Sample
40:12
Complementary DNA (cDNA) Synthesis
41:18
Complementary Synthesis
41:19
Isolate mRNA from Total RNA
41:59
Quantitative PCR (qPCR)
44:14
Technique for Quantifying the Amount of cDNA and mRNA Transcriptions
44:29
Measure of Gene Expression
44:56
Illustration of Read Out of qPCR Machine
45:23
Analysis of the Transcriptome-Micrarrays
46:15
Collection of All Transcripts in the Cell
46:16
Microarrays
46:35
Each Spot Represents a Gene
47:20
RNA Sequencing
49:25
DNA Sequencing
50:08
Sanger Sequencing
50:21
Dideoxynucleotides
50:31
Primer Annealed to a DNA Region of Interest
51:50
Additional Presence of a Small Proportion of a ddNTPs
52:18
Example
52:49
DNA Sequencing Gel
53:13
Four Different Reactions are Performed
53:26
Each Reaction is Run in a Lane of a Denaturing Polyacrylamide Gel
53:34
Example 5
53:54
High Throughput DNA Sequencing
57:51
Dideoxy Sequencing Reactions Are Carried Out in Large Batches
57:52
Sequencing Reactions are Carried Out All Together in a Single Reaction
58:26
Molecules Separated Based on Size
59:19
DNA Molecules Cross a Laser Light
59:30
Assembling the Sequences
1:00:38
Genomes is Sequenced with 5-10x Coverage
1:00:39
Compare Genomes
1:01:47
Entered Into Database and the Rest is Computational
1:02:02
Overlapping Sequences are Ordered Into Contiguous Sequences
1:02:17
Example 6
1:03:25
Example 7
1:05:27
VII. Ethics of Modern Science
Genome Editing, Synthetic Biology, & the Ethics of Modern Science

45m 6s

Intro
0:00
Lesson Overview
0:47
Genome Editing
1:37
What is Genome Editing
1:43
How It Works
2:03
Four Families of Engineered Nucleases in Use
2:25
Example 1
3:03
Gene Therapy
9:37
Delivery of Nucleic Acids Into a Patient’s Cells a Treatment for Disease
9:38
Timeline of Events
10:30
Example 2
11:03
Gene Therapy
12:37
Ethical Questions
12:38
Genetic Engineering
12:42
Gene Doping
13:10
Synthetic Biology
13:44
Design and Manufacture of Biological Components That Do Not Exist in Nature
13:53
First Synthetic Cell Example
14:12
Ethical Questions
16:16
Stem Cell Biology
18:01
Use Stem Cells to Treat or Prevent Diseases
18:12
Treatment Uses
19:56
Ethical Questions
20:33
Selected Topic of Ethical Debate
21:30
Research Ethics
28:02
Application of Fundamental Ethical Principles
28:07
Examples
28:20
Declaration of Helsinki
28:33
Basic Principles of the Declaration of Helsinki
28:57
Utmost Importance: Respect for the Patient
29:04
Researcher’s Duty is Solely to the Patient or Volunteer
29:32
Incompetent Research Participant
30:09
Right Vs Wrong
30:29
Ethics
32:40
Dolly the Sheep
32:46
Ethical Questions
33:59
Rational Reasoning and Justification
35:08
Example 3
35:17
Example 4
38:00
Questions to Ponder
39:35
How to Answer
40:52
Do Your Own Research
41:00
Difficult for People Outside the Scientific Community
41:42
Many People Disagree Because They Do Not Understand
42:32
Media Cannot Be Expected to Understand Published Scientific Data on Their Own
42:43
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Gene Regulation in Eukaryotes

    Medium, 8 examples, 5 practice questions

  • Eukaryotic gene regulation is much more complicated than in prokaryotes, utilizing more regulators and larger regulatory sequences.
  • Signal transduction allows extracellular molecules to cause a chain of events that causes a biochemical response inside the cell.
  • The RNA-induced silencing complex (RISC) is a ribonucleoprotein complex whose most well studied function is the degradation of target mRNA, which decreases the level of transcripts available to be translated.
  • Translational regulation can be specific or global.
  • Epigenetics concerns the inheritance of patterns of gene regulation not found in the DNA sequence itself.

Gene Regulation in Eukaryotes

What is the name of the process where an extracellular signaling molecule activates a specific receptor located on the cell surface or inside the cell?
  • Transcriptional regulation
  • Translation-dependent regulation
  • Signal transduction
  • Epigenetics
Proteins found within cells that are responsible for sensing steroid and thyroid hormones and certain other molecules are called:
  • Ligands
  • Nuclear receptor proteins
  • Polymerases
  • Hormones
What process is very important for gene silencing and viral infection defense?
  • JAK/STAT pathway
  • RNA interference
  • Signal transduction
  • Epigenetics
A type of translation-dependent regulation that rescues ribosomes that are translating mRNAs lacking a STOP codon is called:
  • Nonsense-mediated mRNA decay
  • Nonstop-mediated mRNA decay
  • Signal transduction
  • RNA interference
Methylation of DNA and nucleosomes being passed from generation to generation is an example of:
  • Genetics
  • Molecular Biology
  • Epigenetics
  • DNA replication

*These practice questions are only helpful when you work on them offline on a piece of paper and then use the solution steps function to check your answer.

Answer

Gene Regulation in Eukaryotes

Lecture Slides are screen-captured images of important points in the lecture. Students can download and print out these lecture slide images to do practice problems as well as take notes while watching the lecture.

  1. Intro
    • Lesson Overview
      • Eukaryotic Transcriptional Regulations
      • Example 1
        • Locus Control Regions
        • (Ligand) Signal Transduction
        • N F Kappa B
        • Example 2
          • JAK/ STAT Pathway
          • Example 3
            • Seven-Spanner Receptors
            • Example 4
              • Nuclear Receptor Proteins (NRPs)
              • RNA Interference
              • RISC Pathway
              • Translational Regulation
              • Translation-Dependent Regulation
              • Epigenetics
              • DNA Methylation
              • DNA Methylation
              • Example 5
                • Nucleosomes
                • Chromosome Condensation Via J1
                • Histone Code
                • Example 6
                  • Nucleosome Assembly
                  • Example 7
                    • Chromatin Remodeling
                      • Example 8
                      • Polycomb Repressors
                      • Intro 0:00
                      • Lesson Overview 0:06
                      • Eukaryotic Transcriptional Regulations 0:18
                        • Transcription Factors
                        • Insulator Protein
                      • Example 1 1:44
                      • Locus Control Regions 4:00
                        • Illustration
                        • Long Range Regulatory Elements That Enhance Expressions of Linked Genes
                        • Allows Order Transcription of Downstream Genes
                      • (Ligand) Signal Transduction 8:12
                        • Occurs When an Extracellular Signaling Molecule Activates a Specific Receptor Located on the Cell
                        • Examples
                      • N F Kappa B 10:01
                        • Dimeric Protein That Controls Transcription
                        • Ligands
                      • Example 2 11:04
                      • JAK/ STAT Pathway 13:19
                        • Turned on by a Cytokine
                        • What is JAK
                        • What is STAT
                        • Illustration
                      • Example 3 17:00
                      • Seven-Spanner Receptors 20:49
                        • Illustration: What Is It
                        • Ligand Binding That Is Activating a Process
                        • How This Happens
                      • Example 4 24:23
                      • Nuclear Receptor Proteins (NRPs) 28:45
                        • Sense Steroid and Thyroid Hormones
                        • Steroid Hormones Bind Cytoplasmic NRP Homodimer
                        • Hormone Binds NRP Heterodimers Already Present in the Nucleus
                        • Unbound Heterodimeric NRPs Can Cause Deacetylation of Lysines of Histone Tails
                      • RNA Interference 32:01
                        • RNA Induced Silencing Complex (RISC)
                        • RNAi
                      • RISC Pathway 34:34
                        • Activated RISC Complex
                        • Process
                        • Example
                      • Translational Regulation 41:17
                        • Global Regulation
                        • Competitive Binding of 5 Prime CAP of mRNA
                      • Translation-Dependent Regulation 44:56
                        • Nonsense Mediated mRNA Decay
                        • Nonstop Mediated mRNA Decay
                      • Epigenetics 48:53
                        • Inherited Patterns of Gene Expression Resulting from Chromatin Alteration
                        • Three Ways to Happen
                        • DNA Sequence Does Not Act Alone in Passing Genetic Information to Future Generations
                      • DNA Methylation 50:57
                        • Occurs at CpG Sites Via DNA Methyltransferase Enzymes
                        • CpG Islands Are Regions with a High Frequency of CpG Sites
                        • Methylation of Multiple CpG Sites Silence Nearby Gene Transcription
                      • DNA Methylation 53:46
                        • Pattern Can Be Passed to Daughter Cells
                        • Prevents SP1 Transcription Factors From Binding to CpG Island
                        • MECP2
                      • Example 5 55:27
                      • Nucleosomes 56:48
                        • Histone Core
                        • Histone Protein
                      • Chromosome Condensation Via J1 57:32
                        • Linker Histone H1
                        • Compact DNA
                      • Histone Code 57:54
                        • Post-translational Modifications of N-Terminal Histone Tails is Part of the Epigenetic Code
                        • Phosphorylation, Acetylation, Methylation, Ubiquitination
                      • Example 6 58:52
                      • Nucleosome Assembly 59:13
                        • Duplication of DNA Requires Duplication of Histones by New Protein Synthesis
                        • Old Histones are Recycled
                        • Parental H3-H4 Tetramers
                      • Example 7 1:00:05
                      • Chromatin Remodeling 1:01:48
                      • Example 8 1:02:36
                        • Transcriptionally Repressed State
                        • Acetylation of Histones
                      • Polycomb Repressors 1:03:19
                        • PRC2 Protein Complex
                        • PRC1 Protein Complex
                        • MLL Protein Complex

                      Transcription: Gene Regulation in Eukaryotes

                      Hello, and welcome back to www.educator.com.0000

                      Today's lesson will be on gene regulation in eukaryotes.0003

                      First, we will talk about transcriptional regulation and we will also talk about the regulation of translation.0008

                      Finally, we will talk just a little bit on epigenetics.0013

                      Let us first talk about our eukaryotic transcriptional regulators.0020

                      Mainly, we are going to talk about our transcription factors.0025

                      There are many different types of transcription factors, usually characterized by the specific domain by which they bind DNA.0028

                      We have homeodomain proteins, we have zinc finger proteins, we have leucine zippers, loop helix proteins, as well as HMG proteins.0037

                      HMG just stands for high mobility group.0052

                      We also have insulator proteins, in which these are proteins that bind between the enhancer and the promoter in a gene.0055

                      They suppress transcription.0065

                      If you remember, enhancers are usually far upstream of where the core gene element itself starts.0066

                      Up here, our transcription factors, they can be either enhancing or repressive.0076

                      Compared with our prokaryotes, transcriptional regulation and regulation in general, for eukaryotes is much more complex.0083

                      Usually, there are more regulators and longer length of regulatory sequence in the DNA.0094

                      Let us first start off with an example of transcriptional regulation.0106

                      We have a gal-4 and gal-1.0110

                      Gal-4 is a protein and gal-1 is a DNA sequence.0113

                      We are looking at this in the yeast saccharomyces cerevisiae.0118

                      The gal-4 protein will bind to a site upstream of the gal-1 gene.0125

                      This is going to increase transcription by gal-1 gene, by a thousand fold.0138

                      What this would look like, gal-4 binds as a dimer.0145

                      What we see here, if we draw this out.0150

                      Right here, this right here, this arrow, that shorthand form of saying that is where promoter is and gal-1 that is our gene.0170

                      This right here is a UAS, when upstream activating sequence.0179

                      There are 4 sites here at which the gal-4 protein will bind.0186

                      It will bind as a dimer.0198

                      Between the UAS and the gal-1 gene, is about 235 base pairs.0200

                      The binding upstream over here will affect how this gene is transcribed.0213

                      We can increase, this is gal-4, there are g4.0221

                      This can cause an increase of transcription about a thousand fold.0226

                      For example you are supposed to only make one mRNA0231

                      or in a certain period of time you make a thousand times more than that.0235

                      Let us talk about our locus control regions.0242

                      First off, we are going to draw this out.0245

                      I guess we will use red.0251

                      Here is our locus control region.0256

                      Then, we have our globin genes.0267

                      That should be gamma not Δ.0294

                      This is Δ, this one is β.0307

                      Here is our cluster of globin genes.0320

                      We have locus control regions right.0340

                      Locus control regions are just long range regulatory elements that enhance expression of our linked genes.0342

                      This helps function in a copy number dependent manner.0353

                      It is actually tissue specific.0360

                      For this one, for example, this allows this locus control region.0362

                      This allows ordered transcription of downstream genes.0369

                      What that is really saying is that we have selective expressions of our β globin genes in our erithrocyte cells.0375

                      This is an example talking about our selective.0406

                      Our erithrocyte cells are also known as red blood cells.0413

                      Our locus control regions allow for order transcription in downstream genes.0416

                      Here is our locus control region.0421

                      Here are downstream genes.0425

                      We have our 5 globin genes.0427

                      We have ε, we have gamma G, gamma A, Δ and β.0429

                      During human development, all 5 genes are transcribed sequentially, meaning, first ε, all way through to β.0433

                      What happens here is that we have euchromatin position.0442

                      If we remember correctly, euchromatin is open, more transcriptionally available.0447

                      Heterochromatin is closed, less transcriptionally available.0452

                      Euchromatin position will change and heterochromatin formation, reformation will follow behind.0456

                      This is something that I want you to just keep in mind because we will be talking about epigenetics later.0463

                      How euchromatin and heterochromatin positioning affects the transcription of genes,0472

                      as well as the replication of DNA sequence.0480

                      Here are locus control regions, this is one example.0488

                      Let us move on to signal transduction.0494

                      Signal transduction occurs when we have an extracellular signaling molecule,0498

                      activating a specific receptor located on the membrane of the cell.0506

                      If I draw it out, here is a cell and here might be a receptor.0514

                      Here is our extra cellular signaling molecule.0524

                      When this molecule binds the receptor, it is going to trigger a chain of events, inside the cell it creates some sort of response.0529

                      That response is going to be different, based on which pathway is being activated.0544

                      Some of our examples that we will talk about today are NF kappa B,0550

                      the JAK/ STAT pathway which is very important for immune system regulation.0555

                      We have sigma molecule growth factors, seven-spanner receptors which are important for sensing light, smells, serotonin.0559

                      And then, we have nuclear receptor proteins that are important for the signaling of our steroid and non-steroid molecules.0572

                      This is an important one that you will learn more in biochemistry.0590

                      If you have not taken that yet or if you decide to take a nutrition class, something like that.0594

                      First off, NF kappa B.0603

                      NF kappa B is a dimer protein meaning it has two parts.0607

                      We have to have two pieces of that protein.0612

                      This protein helps control transcription, as well as cytochime production in cell survival.0617

                      Cytochimes are just small protein molecules that are involved in immune system regulation.0622

                      Ligands which means it is just something that is binding, can cause our NF kappa B0628

                      to turn on more than 150 genes in the inflammatory system and the immune system,0637

                      as well as during development in the cytoplasm.0642

                      I mentioned that the NF kappa B is a dimer.0646

                      That dimer is made up of the P50 and the P60 protein, therefore, it is a heterodimer.0650

                      This might be 50, this would be 65, then we have our heterodimer.0657

                      Let us talk a little bit more about NF kappa B.0662

                      NF kappa B, as I said, plays a large part in the immune system and so forth.0667

                      NF kappa B, for one example, can be inhibited when the P50 subunit binds to a cancer suppressor protein.0672

                      If it is inhibited, therefore, it cannot help with transcription or increase transcription, for the rest of its purposes.0686

                      A second example of how NF kappa B maybe regulated, it can be inhibited by its binding to another protein called I kappa B.0696

                      NF kappa B is bound or it can be bound and inhibited by the inhibitory protein I kappa B.0710

                      What you can do to release this inhibition is phosphorylate I kappa B, that will cause I kappa B to release NF kappa B.0720

                      At this point, NF kappa B can travel to the nucleus, it was originally in the cytoplasm.0734

                      At that point, once NF kappa B is in the nucleus, it can go ahead and regulate any transcription, as it normally would.0743

                      At the same time, this phosphorylated I kappa B is no longer bound to NF kappa B, can be ubiquitinated.0752

                      Ubiquity, if we remember back to the protein unit, is just a small protein and that can have many different outcomes.0761

                      In certain outcomes, when a protein is ubiquitinated, it is set to the prodiuzome and degraded,0771

                      broken down, thrown into the trash.0778

                      These are two separate examples how NF kappa B can be regulated, in which case,0781

                      if it is inhibited, it cannot affect transcription.0785

                      If it is not inhibited and able to act in the nucleus, then it can affect and be a regulatory protein.0791

                      Next, we have the JAK/ STAT pathway.0801

                      This pathway is turned on by a cytochime.0803

                      For an example, interferon α, that will attach to the cell surface receptors.0807

                      First of all, let us write out what is JAK.0812

                      JAK is a kinase, it is called Janus kinase.0823

                      Remember, kinases are responsible for phosphorylating things.0832

                      We have STAT, STAT stands for signal transducer and activator of transcription.0837

                      The JAK/ STAT pathway, if we are talking about the previous slide.0878

                      If we have our cell, here is our nucleus.0887

                      We have a receptor and we have something binding to that receptor.0893

                      In this case, the cytokine is binding.0902

                      JAKs, the kinases come together.0906

                      Those JAKs will activate the kinase which phosporylates pairs of our STAT.0911

                      Those phosphorylated STAT monomers will dimerize.0919

                      The stats are also monomers.0923

                      They will be phosphorylated.0933

                      We will just say that it is a yellow piece, that causes them to dimerize.0934

                      They do that via a specific domain on the STAT molecule called an SH2 domain.0954

                      We will call the nucleus in purple over here.0970

                      That STAT dimer will travel into the nucleus and bind a specific DNA sequence, related to transcriptional enhancement.0975

                      This, once again, shows you that a molecule from outside the cell can affect what happens inside the cell,0987

                      through a series of events that happens to, in this case, deal with the binding of a molecule to receptor,0995

                      a phosphorylation of other proteins.1006

                      Those proteins being sent into the DNA or being sent into the nucleus, to bind to DNA.1009

                      At that point, it will affect transcription.1016

                      Another example of sigma transduction would be our signal molecule growth factor.1022

                      A couple examples being epidermal growth factor or EGF, as well as insulin.1028

                      Remember, insulin is the protein that is sent out by the body, by the pancreas, in response to glucose in the blood, blood sugar.1033

                      When you eat something, you have carbs, those get broken down, absorbed from the stomach through the intestine into the blood.1046

                      Insulin is released, insulin takes that glucose into the cells.1055

                      How this works is that, we have a binding to the receptor, activating a dimeric, a two protein,1061

                      RTK which is a receptor thyrosine kinase enzyme.1076

                      The dimerization requires the adapter proteins grab to an SOS.1082

                      That is just some extra information for your knowledge.1088

                      We have this signal causing the RTK dimerization.1092

                      That dimer binds and activates a protein called RAS, which will bind the GTP molecule.1102

                      That RAS will then bind to and activate another protein called RAF.1110

                      RAF is what is called a map KKK which is monogen activated protein, we usually call this MAP.1115

                      It is a MAP kinase kinase kinase meaning it phosphorylates a MAP kinase kinase.1127

                      It is quite heavy nomenclature but we will understand it, as we go to the next step.1138

                      This RAF which is our enzyme, a specific MAP kinase kinase kinase.1143

                      RAF will then phosphorylate, since it is a kinase, it will phosphorylate MEK.1149

                      MEK is a MAP kinase kinase.1155

                      MEK, once it is phosphorylated, will phosphorylate a MAP kinase, a MAP K.1162

                      This MAP K is a different type of kinase, than the one that we talked about up here,1175

                      it is a kinase that is called a serine threonine kinase.1182

                      What are the differences between these kinases?1187

                      The RTKs, they phosphorylate a thyrosine amino acid.1189

                      This one down here will phosphorylate proteins at certain amino acids of either a serine or a threonine residue.1199

                      Now that MAP kinase is active, it is still a kinase.1213

                      MAP kinase will phosphorylate a bunch of other transcription factors.1218

                      These transcription factors, now many of them by the phosphorylate, will be activated.1224

                      And now, they can go and bind to the DNA in the nucleus and enhance transcription.1231

                      It is a very complex process, many steps, but the whole outcome is to affect the transcription happening in the nucleus.1239

                      We have other ways of regulating transcription.1252

                      Utilizing certain proteins called seven-spanner receptors.1257

                      What is a seven-spanner receptor?1262

                      If this is the cell membrane, we have a protein that crosses the cell membrane 7 times, 1, 2, 3, 4, 5, 6, 7.1266

                      Maybe that is its N terminal tail and this is its C terminal tail.1284

                      What we have here is it is crossed 7 times, 1, 2, 3, 4, 5, 6, 7.1288

                      That is what is a seven-spanner receptor is.1296

                      There are several different seven-spanner receptors.1299

                      How do these work?1303

                      Very similar, in the fact that there is ligand binding that is activating a process.1306

                      Specifically, we have ligand binding, activating trimeric proteins called G proteins.1313

                      Trimeric meaning 3, 3 different polypeptide chains.1320

                      Each one of these subunits, we have an α subunit, a β subunit, and a gamma subunit.1328

                      In order, how this happens, we have the ligand binding activating that protein.1339

                      When GTP binds, our α subunit that will activate adenylyl cyclase, which is an enzyme.1347

                      This requires GTPAs activating protein also known as GAP.1363

                      Adenylyl cyclase normally converts ATP to cyclic AMP.1372

                      We talked about this in the previous lesson, this adenylyl cyclase enzyme.1376

                      The CAMP, we have also talked about this, will bind to protein kinase A to activate it.1383

                      Protein kinase A, otherwise known as PKA, will then be able to phosphorylate many different proteins.1391

                      Some of which can be transcription factor.1401

                      PKA, once activated can phosphorylate transcription factors which will bind to certain pieces of DNA in the nucleus and affect transcription.1403

                      Protein kinase A can also another protein called CREB which will bind to a specific DNA sequence called CRE.1416

                      As well as, be bound by CBP and P300 protein which is CBP stands for creb binding protein, CREB BP.1429

                      CBP300, this protein is normally considered a transcriptional activator.1444

                      It turns on the transcription of genes.1450

                      There are two different ways a protein kinase can affect gene expression.1453

                      Let us talk about another example.1465

                      We actually have two examples in this one.1467

                      Let us talk about how we have two different diseases, cholera and whooping cough, otherwise known as pertussis.1470

                      Cholera, cholera is caused by the action of the bacteria vibrio cholerae.1479

                      This vibrio cholerae produces a toxin called the cholera toxin.1495

                      How does this work, what is the mechanism of action?1500

                      The cholera toxin catalyzes an ADP ribosylation.1504

                      That is basically adding a ribose group to ADP together.1509

                      This happens via the use of our old friend NAD, nicotinamide adenine dinucleotide.1514

                      We have talked about this one before and it is useful in energy.1523

                      This ATP ribose will bind to the α subunit of the G protein, as we talked about in previous slide,1529

                      which does not allow the GDPAs activating protein, GAP, to activate GTPAs.1538

                      Therefore, you cannot convert GTP to GDP.1545

                      What is that even mean?1551

                      What that means that we have constitutive activation of our adenylyl cyclase enzyme.1553

                      Constitutive means it is always on.1562

                      If our adenylyl cyclase enzyme is always on, that means that we are continually turning ATP into CAMP.1566

                      If we have a bunch of CAMP, that is going to keep PKA, protein kinase A, constitutively active.1577

                      If PKA is always active, it is always going to be phosphorylating things.1587

                      In these certain cases, it will phosphorylate certain transcription factors to turn on gene expression and1594

                      can lead to types of diseases or the types of symptoms from cholera or whooping cough that are common.1601

                      Whooping cough, specifically, is caused by the bacteria bordetella pertussis and1614

                      that bacteria also produces a toxin called the pertussis toxin.1621

                      This pertussis toxin acts in a little different way than the cholera toxin does.1625

                      Let us talk about this, we have pertussis toxin binding to a protein called G sub I.1632

                      G sub I is a G protein inhibitor.1640

                      Normally, G sub I will bind to and inhibit adenylyl cyclase, opposite of cholera.1647

                      Cholera adenylyl cyclase is always on.1660

                      G sub I normally turn adenylyl cyclase off.1663

                      However, we have pertussis toxin binding to that inhibitor preventing that normal inhibition.1667

                      Therefore, we end up with the same outcome.1680

                      We have constitutive activation of adenylyl cyclase, leading to continual conversion of ATP to cyclic AMP.1683

                      Meaning, continual activation or constitutive activation of protein kinase A,1694

                      leading to transcriptional regulation at increased rates.1701

                      These are just a couple of ways how we can utilize the previous material in this unit,1708

                      to understand how a couple different bacteria create their symptoms that we see for medical purposes.1715

                      Going on to a different type of transcriptional regulation.1728

                      We have what are called nuclear receptor proteins.1732

                      These are proteins found within cells that are responsible1736

                      for sensing both steroid and thyroid hormones, as well as certain other molecules.1739

                      First off, we have our steroid hormones.1749

                      For example, our androgens and estrogens, our sex hormones, our glucocorticoids which are involved in water retention.1752

                      They are going to bind our cytoplasmic nuclear receptor protein homodimer.1761

                      Let us say dimer of the same two things, two different polypeptides.1768

                      The steroid hormones, these will bind our NRP homodimer.1774

                      This is what is called a type 1 complex.1782

                      The type 1 complex will head the nucleus, bind to an inverted repeat DNA sequence called a hormone response element or an HRE.1787

                      At that point, once the HRE is bound then transcription can be regulated, either in a positive or negative manner.1802

                      A second way that nuclear receptor proteins act is sensing hormones, such as vitamin A or a thyroid hormone.1811

                      They will bind to NRP heterodimers.1822

                      In this case, it is two different proteins.1826

                      This is what is called a type 2 complex.1836

                      This type 2 complex is already in the nucleus.1840

                      It will bind direct repeats in the nuclear DNA and then affect transcription regulation.1845

                      A third way to look at our NRPS and how they might affect transcription1855

                      would be in an unbound heterodimic NRP, not bound by hormone.1862

                      They can cause deacetylation of histone tails, the lysine residues on histone tails.1868

                      These heterodimeric NRPs that are bound by a hormone, such as this right here, can cause the acetylation of those tails.1876

                      Remember, when a histone, when its tails are acetylated, depending on the histone code, it is very complex.1887

                      It often is related to the loosening of chromatin meaning transcriptional activation.1899

                      Whereas, the deacetylation of histone tails usually means the compaction of chromatin,1908

                      meaning decreased transcription and replication.1915

                      Let us move on to something that is also regulating transcription but in a completely different way.1923

                      Actually, this is a really cool mechanism.1932

                      This is what is called RNA interference.1937

                      This is a method that is actually been adopted by many research labs.1939

                      I will show you an example in a couple slides to actually choose what genes get silenced or expressed.1945

                      This is coordinated by a complex called the RISC complex.1958

                      The RNA induced silencing complex.1964

                      It is a ribonuclear protein complex meaning it has both RNA and protein.1970

                      This complex has been most well studied for having a function in the degradation of a specific target mRNA.1977

                      If you target mRNA to be degraded, that will decrease the level of the transcripts available to be translated.1988

                      Therefore, in S sense it is functioning to decrease gene expression of a specific gene.1996

                      We have very important players.2006

                      We have RNase enzymes, RNase is an enzyme that breaks down RNA molecules.2009

                      These RNase enzymes called dicer and drosha, trim double stranded RNA2016

                      to form either small interfering RNA, sRNA, or microRNA, miRNA.2024

                      There RNAs then get incorporated into the RISC complex leading to specific mRNA targeting, and then degradation.2035

                      It targets to a specific mRNA and does not allow translation to take place.2046

                      It is actually really cool.2052

                      This process we, call RNA interference or RNAi.2055

                      We find this in many eukaryotes.2061

                      It was thought to be developed as a very important mechanism of gene silencing and viral infection defense.2063

                      Let us talk about the RISC pathway, in a little bit of detail.2076

                      First of all, what is an activated RISC complex?2080

                      That composes the RISC ribonuclear protein, as well as a micro RNA or small interfering RNA and an ATP molecule.2083

                      How does this all go down?2096

                      First off, dicer and drosha cleave a double stranded RNA into short 21 to 23 base pair fragments,2100

                      with a two nucleotide 3 prime overhang.2111

                      Simply, it looks like this, where this is two nucleotides, that is two nucleotides long.2115

                      This is our 3 prime, 5 prime, 3 prime, 5 prime.2137

                      This whole thing is 21 to 23 base pairs.2145

                      This is double stranded RNA.2156

                      That is the first thing that happens.2165

                      Then, we bring in a different RNase called argonaut which is also termed slicer.2167

                      The RNase argonaut then associates the single stranded RNA with the RISC complex,2178

                      to act as a complementary strand to whatever the target mRNA is.2190

                      This double stranded piece right here, that has been processed by dicer and drosha2198

                      can be separated into single strands and that is what argonaut is working with.2206

                      The complex will bind to our target mRNA, whatever piece of gene material2210

                      that you do not want to end up turning into a protein.2219

                      The complex binds to the target mRNA and silences it.2223

                      Silencing meaning it is not been turned into a protein, no translation.2226

                      There are two different ways this occurs.2232

                      If we are talking about an miRNA being used, micro RNA.2234

                      The micro RNA in the RISC complex binds to the 3 prime untranslated region,2242

                      that is a part of an RNA, and is untranslated.2249

                      Therefore, it would not have turned into protein anyway2252

                      but is a great spot for regulatory proteins to bind and affect whether something is translated or not.2256

                      MiRNA in that RISC complex, binds to the 3 prime UTR with a mismatch.2262

                      It will block transcription.2270

                      The other way that we can affect translation, if we have a small interfering RNA in the RISC complex,2274

                      binding to the mRNA without a mismatch.2285

                      This will cause RISC to cleave the mRNA.2291

                      Cleaving it vs. just blocking transcription.2296

                      mRNA degradation happens at a certain spot in the cell called a P body.2304

                      mRNA degradations localized in P bodies which are found in cytoplasmic space.2314

                      And often stain darkly, that you can look under a microscope.2321

                      Importantly, we have activated RISC, the RNA inducing complex,2329

                      being implicated in formation of nuclear heterochromatin.2339

                      Meaning, we can have RISC actually not just cleaving these mRNAs but can actually somehow be a part of epigenetic changes.2342

                      Actually not even getting transcription available, not even allowing transcription to happen.2358

                      That is kind of cool.2364

                      Here is an example of the RISC pathway.2369

                      Actually, how scientists have been able to utilize it.2371

                      What we would have is viral, if there is an infection, they make double stranded RNA.2378

                      You can break that into srRNA via dicer and drosha.2386

                      Those double stranded RNAs can enter into the RISC complex, argonaut can take a single stranded RNA2391

                      and utilize that either as an miRNA or srRNA, to either degrade the mRNA2398

                      or to affect possibly the epigenetic state of the heterochromatin.2406

                      What scientists can do, they can target individual genes and silence them.2414

                      Here we see a wild type, a normal petunia plant.2420

                      These two are lab created petunias, utilizing RNA interference, RNAi, using the RISC pathway.2426

                      What they have done is through selective targeting of the mRNA specific for the color,2442

                      they have silenced them in the white portions.2448

                      This is pretty cool.2453

                      RNAi can be used other than just for cool phenotypes.2455

                      It can actually possibly be used for more functional things such as something that we will talk about a little later,2464

                      maybe gene therapies, something similar to that.2471

                      That is your transcriptional regulation.2479

                      We will just have a few slides, a few instances of translational regulation.2482

                      Just like prokaryotes, eukaryotes mostly regulate transcriptionally.2486

                      However, they can regulate translationally.2494

                      You can have global regulation of translation meaning you regulate all translation,2498

                      not specific genes, by phosphorylating protein EIF2 that we talked about before.2505

                      This is eukaryotic initiation factor 2.2515

                      If you phosphorylate EIF2 that protein cannot bind GTP.2519

                      If you cannot bind GTP, you are not able to bring the starting codon, the charged tRNA with the initiating methionine.2525

                      You cannot bring that to the ribosome because this EIF2 GTP complex is required to bring that to the 40S ribosome.2537

                      If you phosphorylate EIF2, you cannot translate anything.2547

                      Another way of global regulation, we have phosphorylate of EIF2,2554

                      another form of global regulation is competitive binding of the 5 prime cap of mRNA.2559

                      Normally, that mRNA, remember, we have a cap.2566

                      That cap is normally bound by EIF4-G.2572

                      Phosphorylated regulatory proteins called 4EBP, binding protein, the cap is normally bound by EIF4-E which is then bound by EIF4-G.2583

                      If these EIF4-E is bound by these 4E binding proteins right, then the EIF4-E will not allow,2614

                      the binding of 4EBP will not allow EIF4-G to bind.2669

                      Therefore, you will not allow translation to occur.2673

                      This can also be used to regulate specific mRNAs but in general, if you affect the binding of initiation factors for translation,2679

                      binding the actual mRNA you can affect all mRNAs that are undergoing translation.2690

                      We can talk about a couple different ways of translational regulation.2698

                      This is what is called translation dependent regulation.2703

                      This is when translation is already started and it is in the elongation phase.2706

                      We have two different types, we have nonsense mediated mRNA decay and we have nonstop mediated mRNA decay.2712

                      Nonsense mediated mRNA decay is when you come across an mRNA that has a premature stop codon.2723

                      What you want to do is degrade those mRNAs so that you do not even have to utilize the energy and2734

                      the amino acids of the whole process to make this short protein which will end up being degraded by the prodiuzome anyway.2741

                      You can degrade mRNAs with the premature stop codon, by removing either the 5 prime cap2751

                      or the 3 prime poly-A tail of the mRNA.2760

                      In which case, you have now freed up either one of those ends to nucleolytic digestion.2764

                      Exonucleases can now come in and just chew it up and degrade it, the mRNA.2771

                      We have another thing called nonstop mediated mRNA decay.2777

                      This occurs when a ribosome is working on an mRNA that wacks a stop codon.2781

                      This can be pretty detrimental to the cell because if there is no stop codon,2789

                      basically the ribosome is stuck with that mRNA attached to it, as well as the protein coming out.2794

                      It just stalled, nothing can happen, you cannot release the polypeptide.2812

                      You cannot separate the ribosome, which means you cannot then translate a new mRNA.2817

                      Without that stop codon, the ribosome is stalled, it cannot do anything.2825

                      We want to be able to fix that.2833

                      This is rescuing, ribosome is translating mRNAs without a stop.2836

                      If you do not have a stop codon, that poly-A sequence in the tail gets translated,2842

                      and that Poly-A, the AAA codon gives you lysine.2849

                      What this does, that causes the ribosome to stall.2854

                      It acts as a signal that there is something wrong.2858

                      What happens is we have eukaryotic releasing factors 1 and 3,2862

                      binding to the ribosome and associating the ribosome from the mRNA.2869

                      At that point, the endonuclease which actually is an unknown endonuclease at this point,2892

                      researchers are still dwelling into this information.2899

                      This endonuclease will degrade the mRNA from 3 prime to 5 prime.2902

                      The protein, if it is enable to fold properly which is very likely due to all of the extra lysines,2908

                      will likely just be signaled to be degraded and be sent to the prodiuzome.2917

                      These are two different ways that you can have translational regulation, actually once translation has occurred.2922

                      Transcriptional regulation and translational regulation, and now I’m on to epigenetics.2936

                      We touched over this a little bit, when we talked about chromatin organization.2942

                      I think it was unit 5 but we are going to talk a little bit about this again, give you a little more detail.2946

                      Epigenetics, what is it?2956

                      That is inherited patterns of gene expression resulting from chromatin alteration.2958

                      It is not in alteration of the DNA sequence that is heritable.2964

                      It is not a change in the base sequence, A, C, T, G.2969

                      But it is a heritable, passing from generation to generation,2974

                      it is a heritable trait of extra non DNA related changes, in terms of the basic cell.2980

                      But it could be due to just small changes, such as a methylation, such as acetylation.2996

                      It can be on the DNA sequence, as well as on the nucleusome itself.3004

                      This can happen, we will talk about three different ways, DNA methylation,3017

                      the nucleosomes or the histone proteins, as well as something called polycomb repressors.3023

                      A very important point of epigenetics is that DNA sequence does not act alone in passing genetic information to future generations.3028

                      Meaning, the methylation state, let us say of DNA or of histone proteins.3039

                      The methylation state usually has to do with what genes are silenced or active, that is able to be sent from generation to generation.3046

                      DNA methylation usually occurs at CPG sites or CG sites.3059

                      The p that just says it is a C right next to a G, only separated by a phosphate which is what is in the backbone of DNA.3064

                      DNA methylation occurs in our CPG sites via the DNA methyltransferase enzyme.3073

                      What you do is you methylate cytosine to get 5-methylcytosine.3081

                      That can keep that methylation even through the next generation.3086

                      However, one drawback to this is that actually spontaneous deamination can turn this 5-methylcytosine into thymine overtime.3092

                      Remember, this cytosine, even as a 5-methylcytosine, will base pair with a guanine.3103

                      If it gets deaminated, it now becomes a thymine.3113

                      On the other strand that was not touched, it is a guanine.3120

                      What you need to do now is, we see that this is in a proper DNA pair, there is a DNA repair that needs to come on.3124

                      If this T is repaired back to a C, no harm no foul.3133

                      If this T is not repaired but the G is repaired to an A, now we have a difference in the original base sequence.3145

                      An epigenetic change has actually turned into a genetic change that gets passed on to next generation.3157

                      This can cause DNA mutations, if not properly repaired.3167

                      CPG islands, I just want to talk about because you will hear that a lot, probably,3173

                      if you read some research papers or if your professors are talking about genetics and especially DNA methylation.3177

                      CPG islands are regions of DNA with the high frequency of CG sites.3184

                      There are usually about 200 base pairs in length and they are associated with promoter regions of a lot of our mammalian genes.3191

                      Usually, when you see CPG island, that is something that should bring to your mind, we are in the promoter of a gene.3199

                      Not all the time, but very frequently.3209

                      Methylation of many of these CPG sites will silence gene transcription,3213

                      leading to heterochromatinization, the formation of heterochromatin.3219

                      The methylation pattern, as I talked about before, it can be passed on to daughter cells.3228

                      This methylation will prevent the binding of a transcription factor called SP1 to the CPG islands.3233

                      The binding of SP1 is part of transcriptional enhancement.3242

                      We also have this other protein involved with DNA methylation called MECP2.3250

                      This is a binding protein of those methylated CPG islands.3257

                      It is a repressor protein that binds to those islands in DNA.3263

                      It will act as a transcriptional repressor.3268

                      It decreases transcription by recruiting histone deacetylase.3272

                      Remember, if you deacetylate histones that usually compacts our chromatin,3275

                      meaning there is no room for transcription of machinery to get in and go through making your mRNA.3281

                      A phosphorylated MECP2 protein has a decreased infinity for methylated CPG sites, and not only to an increase in transcription.3291

                      MECP2 repressor protein, phosphorylate it, it would not bind, therefore, it leads to transcriptional activation.3309

                      SP1 transcription factor, that is an activator protein.3317

                      Let us look at an example using MECP2.3324

                      We have a protein called BDNF, brain derived neurotrophic factor.3329

                      It is a protein implicated in learning a memory class disease.3335

                      Basically, being able to retain your memories and reorganize them.3338

                      This BDNF is release when our dendrites which is a neuro cell, get depolarized, activated.3344

                      BDNF release can lead to a phosphorylation of in MECP2, meaning, it will now not be able to bind the DNA very well,3351

                      meaning you have an increase in transcription.3362

                      On a related but different specifically from this BDNF, if you have a mutation in the gene for the MECP2 protein3368

                      which is also the MECP2 gene, this will cause a disease called rett syndrome.3378

                      This is a syndrome that has clinical manifestations very similar to autism.3385

                      It will affect the neural function.3390

                      In addition to the autism like symptoms, you have stereotypical hand movements3393

                      like handwringing or repeatedly putting your hands in your mouth.3399

                      It is almost unconscious.3403

                      These are some slides that we have already seen in the chromatin organization lecture.3410

                      But I wanted to just have them again so that we can reference them.3416

                      Remember, we a histone core, we have 8 histone proteins, we have 2 dimers of H2 and H2B,3420

                      one tetramer of H3 and H4.3429

                      147 base pairs of DNA wound around.3433

                      It is about 1.65 times around the core.3437

                      We also have linker DNA, 20 to 60 base pairs.3440

                      Just that nucleosome will compact DNA about 6 fold.3446

                      When you add in the linker histone H1, you can compact even more into your 30 nm fiber from your 10nm,3452

                      that will compact DNA about 40 fold.3462

                      You can even compact even more than that, utilizing scaffolding so that you can make a bunch of loops.3466

                      We have talked about the histone code before, there are post translational modifications of our N-terminal histone tails.3477

                      That is part of the epigenetic code.3485

                      If we remember, phosphorylation usually adding a negative charge, acetylation positive charge.3487

                      If you are adding a negative charge that is going to decrease its interaction with DNA.3494

                      It will loosen it, making it more transcriptionally active.3502

                      Acetylation adding positive charge.3506

                      That is neutralizing the negatively charged lysine which is actually going to open you up.3513

                      Methylation will increase the interaction of DNA meaning transcriptional repression.3519

                      You also have ubiquitination which can be binding site for either transciptional activators or repressors.3524

                      We have talked about the histone code being very complicated.3534

                      You can silence, you can have gene expression, gene silencing.3537

                      You can have chromosome condensation, you can have DNA damage repair,3541

                      based on several slight changes in the epigenetic code of the histone.3547

                      We talked about nucleusome assembly.3556

                      When you replicate your DNA, you have to replicate your nucleosomes.3558

                      Some of those nucleosomes get reused in the next generation.3563

                      What those nucleosomes, whatever they had for epigenetic changes, methylations, ubiquitinations, phosphorylations,3569

                      acetylation, whatever, that will be transported with the nucleosome to the daughter cell.3578

                      That can propagate the epigenetic status, from one generation to the next.3587

                      What is very important are those parental H3 H4 tetromers.3600

                      We have talked about the inheritance of the chromatin state.3606

                      Histone acetlytransferases combined acetylated tails using its bromo domain.3611

                      Therefore, spread the acetylation which would decrease the condensation of the heterochromatin,3617

                      to make it more euchromatic, therefore, transcriptionally active.3631

                      Or you can have the same thing happening with histone methyltransferases3636

                      binding methylated regions via its chromo domain, causing a condensation.3641

                      Therefore, more heterochromatization IE transcriptional repression.3649

                      We will just write this real quick.3657

                      Histone acetyltransferase binds acetyl groups via its bromo domain leading to a looser state3660

                      which is more likely going to be a transcriptional activation.3679

                      We have histone methyltransferase via its chromo domain, binding methylated histone tails3685

                      which will tighten, condense, leading to a more likely repressed state.3698

                      Chromatin remodeling, it is all part of epigenetics.3711

                      As I said on the previous slide, when we have histone acetyltransferase,3715

                      as well as chromatin remodelers like the swi/snf complex,3718

                      that allows you to loosen up the interaction of DNA with the histones and allow your transcription of machinery to come in.3722

                      Therefore, being in an activated state.3732

                      If you have your histone deacetylases and your histone methyltransferases, that is going to cause a tighter association with DNA.3735

                      Therefore, less room for your big RNA polymerase and all your initiation factors to come in and allow transcription.3743

                      This is a transcriptionally repressed state.3751

                      Our last example, I believe, we have methylation of DNA and histones seeming to be correlated.3757

                      Usually, when both DNA and histone tails are methylated, this will be our transcriptionally repressed state.3765

                      Acetylation of histones leads to loosening of DNA around the histone, offering a high likelihood of replication or transcription.3773

                      Acetylation, high likelihood of replication and transcription.3784

                      Methylation, transcriptionally repressed state.3792

                      Finally, just as a mentioned, we have our polycomb repressors.3801

                      These polychrome repressors just are a family of proteins that can remodel chromatin,3806

                      such that epigenetic silencing will take place.3813

                      PRC, that is just polycomb repressor complex or repressive complex.3818

                      PRC2 protein complex will bind DNA and try methylate your histone 3 at lysine 27.3825

                      This will cause a repression.3834

                      The PRC1 protein complex will also repress transcription.3841

                      We have another protein complex called the MLL protein complex which will reverse this repression by demethylating H3K27.3846

                      A couple things I want to point out is that, the PRC1 protein complex, a mutation in this,3859

                      if you are a mutant, like they can make in labs, mutants die perinatally.3870

                      Meaning, either at the time of birth or shortly after no more than two weeks, in under a couple of weeks.3879

                      The PRC2 mutants are what are called embryonic lethal, meaning they never get born in the first place.3896

                      These are really important complexes.3913

                      One more thing to show you how important they are is that, over expression, if PRC1 and PRC2 are too active,3915

                      either too many around, they are repressing too much.3925

                      If they are over expressed that is actually correlated with a more severe and more invasive types of cancer.3927

                      Your cancer is likely to be, you are worse off if you have over expression of your polycomb repressor complexes.3942

                      That is the end of the lesson today, I hope you enjoyed our lesson.3952

                      I hope I see you back for our next one.3957

                      Thank you for joining us at www.educator.com, I hope to see you again.3963

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