1. Gene Expression
Dr. Kevin Ahern

2. Gene Expression

3. Controls on Protein Levels
Gene Expression
Controls on Protein Levels
Transcription
Splicing
Polyadenylation
mRNA Stability
Translation
Protein Stability

4. Transcription Control - Prokaryotic Promoter

5. Transcription Control - Prokaryotic Promoter
Polycistronic Message in Prokaryotes

6. Transcription Control - Prokaryotic Promoter
Allo-Lactose
Lactose

7. Transcription Control - Prokaryotic Promoter

8. Transcription Control - Prokaryotic Promoter

9. Transcription Control - Prokaryotic Promoter
CAP
cAMP
CAP Site of DNA

10. Transcription Control - Prokaryotic Promoter
From Wikimedia Commons

11. Prokaryotic Transcription Control - Termination/Attenuation
From Wikimedia Commons

12. Prokaryotic Transcription Control - Termination/Attenuation

13. Transcription/Translation Control - Riboswitches
Cis-acting sequences
Anti-terminator
Terminator

14. Transcription/Translation Control - Riboswitches
Lysine Bound to Riboswitch

15. Eukaryotic Gene Expression
Much More Complexity
Chromatin
Many Transcription Factors
Enhancers

16. Transcriptional Control - Eukaryotes

17. Increasing Magnification
RNA
Eukaryotic Gene Expression - Chromatin
Chromatin is the Complex of DNA, Protein, and RNA Comprising Eukaryotic Chromosomes
For RNA Polymerase to Perform Transcription, Access Must Be Gained to the DNA
Increasing Magnification

18. RNA
Eukaryotic Gene Expression - Epigenetics

19. RNA A Nucleosome is a Fundamental Unit of Chromatin Structure
Eukaryotic Gene Expression - Chromatin
A Nucleosome is a Fundamental Unit of Chromatin Structure
Contains Two Copies Each of Four Histone Proteins - H2a, H2B, H3, and H4
DNA is Wrapped Around this Octet Core and Histone H1 is on the Outside

20. RNA
Eukaryotic Gene Expression - Chromatin
Histone Proteins Are Rich in Basic Amino Acids, Making Them Positively Charged
The Positively Charged Proteins Are Attracted Strongly to the Negatively Charged Phosphates of the DNA
Chemical Modifications That Affect These Charges Influence Transcription

21. RNA
Eukaryotic Gene Expression - Chromatin

22. Chemical Modification
RNA
Eukaryotic Gene Expression - Chromatin
Histone Acetyl Transferases (HATs) Use Acetyl-CoA to Put Acetyl Groups on Lysines in Histones
This Neutralizes Their Positive Charge and Loosens Interactions With the Histones, Facilitating “Remodeling” or Restructuring of Chromatin to Allow Transcription to Occur
Acetylated Lysines Can Also be Binding Targets for Proteins Affecting Transcription
Chemical Modification
Unwinding of Complex

23. RNA
Eukaryotic Gene Expression - Epigenetics

24. Transcription Complex
RNA
Eukaryotic Gene Expression - Chromatin
Histone Acetylation Favors Euchromatin and Stimulates Transcription
Histone De-Acetylases Reverse These Effects, Favoring Heterochromatin and Gene Silencing
The Sirtuin 1 deacetylase in humans down-regulated with insulin resistance
Numerous Chemical Modifications are Made to Histone Proteins
Acetylation / Deacetylation
Methylation / Demethylation
Phosphorylation / Dephosphorylation
Ubiquitination
Chemical Modification to Bases in DNA Can Also Affect Transcription
Open and Accessible to
Transcription Complex
Condensed and
Not Accessible

25. RNA
Eukaryotic Gene Expression - Epigenetics

26. Epigenetics
Chemical Modifications in Histones and DNA Can Cross Generational Barriers
Transcriptional Effects Can Thus Be Transmitted From Parent to Progeny Independent of the Sequence of the DNA.
Such Influences are Called Epigenetic

27. Epigenetics Överkalix study
Patterns of modification of chromosomes cross generational barriers.
Genetic Imprinting
Överkalix study
1. A greater body mass index (BMI) at 9 years in sons, but not daughters, of fathers who began smoking early.
2. The paternal grandfather's food supply was only linked to the mortality RR of grandsons and not granddaughters.
3. The paternal grandmother's food supply was only associated with the granddaughters' mortality risk ratio.
The estimation of the percentage of human genes subject to parental imprinting is approximately one to two percent, currently parental imprinting has been identified in fewer than 100 distinct named genes.

28. Transcriptional Control - Eukaryotes
Methylation of CpG sequences in
eukaryotes inhibits transcription

29. Transcriptional Control - Eukaryotes

31. Transcriptional Control - Eukaryotes

32. RNA Blocking Insulators Allows Enhancer to Activate Transcription
Eukaryotic Gene Expression - Transcription
Blocking Insulators Allows Enhancer to Activate Transcription
Insulators Can Block Enhancer’s Activation of Transcription

33. Iron Transfer & Storage
Ferritin - Cellular Protein to Bind Iron
Transferrin Receptor - Membrane Protein to Transfer Iron

34. Iron Transfer & Storage
Iron Response Element
Binding Protein (IRE-BP)
Iron Response Element (IRE)

35. Iron Transfer & Storage - Translation Regulation

36. Iron Transfer & Storage - mRNA Stability

37. RNA
RNA Interference
RNA Interference is a Powerful Means of Controlling Gene Expression
Viral and Endogenous Cellular Genes Are Targets
A Similar System Called piRNA (piwi RNA) Protects Against Transposon Genes
Considerable Interest in Using Technique to Genetically Transform Organisms for Protect Against Pathogens

38. RNA Cellular Source Cellular Pre-Processing
RNA Interference
Transcription
Double-Stranded RNA is Stimulus
Processing
Viral Infection
20 bp pieces
RISC
RISC
Target Complementary Sequences in mRNAs

39. + + RNA Complementary Sequences Align Argonaute Activity
RNA Interference
Complementary Sequences Align
RISC
RISC +
Argonaute Activity
in RISC Breaks
mRNA, Stops
Translation
mRNA
Translation of mRNA Stopped
RISC +

40. RNA Protection Against Invading Viruses Stimulated by dsRNA
RNA Interference
Protection Against Invading Viruses
Stimulated by dsRNA
miRNA (cellular) & siRNA (foreign)
Cellular piwi RNAs (piRNA) have similar functions in silencing transposons
Widespread in Eukaryotes
Actions referred to as RNA Interference (RNAi)
RNA Interference Operates Through the Silencing of Gene Expression
DS RNA induces Dicer to chop it into 20 BP Pieces
These siRNAs/miRNAs bind to the RNA Induced Silencing Complex (RISC)
One Strand is Destroyed and One Retained to Bind to Complementary mRNA sequences
RISC Nuclease Activity (Argonaute)
1. Destroys mRNA Where Strand Binds or
2. siRNA/miRNA strand on mRNA blocks translation or
3. si/RNA/miRNA strand destabilizes mRNA and Targets for Destruction

41. RNA Bonding to mRNA Premature Stopping of Translation Degradation of
RNA Interference
Bonding to mRNA
Premature
Stopping of
Translation
Degradation of
mRNA