1. Eukaryotic Gene Expression
Managing the Complexities of Controlling Eukaryotic Genes

2. Prokaryotes vs. Eukaryotes
Closely related genes are clustered together
Related genes are located on different chromosomes
Largely transcriptional control
Significant transcriptional control; other levels of control possible
Larger number of larger-sized genes
Trans-acting sequence-specific DNA binding proteins
Proximal Cis-acting sequences
Cis-acting sequences can be located at significant distances

3. Control Points for Gene Expression in Eukaryotes
DNA
transcription
Transcriptional Control
RNA
Post-Transcriptional Control
translation
Translational Control
Protein
Post-Translational Control

4. Levels of Eukaryotic Chromatin Structure
First level of chromatin coiling
Nucleosome = DNA + histone proteins
Variations in chromatin condensation affect gene activity.

5. Transcriptional Regulation: Effects of Chromatin Structure
Decompaction of chromatin:
Transcription factors unwind nucleosomes in the area where transcription will begin, creating DNAse I hypersensitive sites
RNA polymerase unwinds more nucleosomes as transcription proceeds

6. Transcriptional Regulation: Effects of Chromatin Structure
Acetylation of histone proteins (adding -CH3CO) reduces DNA-histone interaction, permitting transcription factors to bind.

7. Transcriptional Regulation: Effects of Chromatin Structure
DNA Methylation
DNA Methylation (adding -CH3) can occur on cytosines at CpG groupings near transcription start sites
Inactive genes have methylated cytosines
Active genes have demethylated cytosines
Acetylation of histones is associated with cytosine demethylation

8. Transcriptional Regulation: Control of Initiation
Transcriptional Activator Proteins assist in the formation or action of the basal transcription apparatus

9. Transcriptional Regulation: Control of Initiation
Transcriptional Activator Proteins bind to Enhancer sequences that increase transcription
Enhancers can influence promoters at distances of 50 kb or greater due to DNA looping mechanism
Insulators control the direction of enhancer action

10. Transcriptional Regulation: Control of Initiation
Transcriptional Repressor Proteins have three possible modes of action
compete with activators for DNA binding sites
bind to sites near activator site and inhibit activator contact with basal transcription apparatus
interfere with assembly of basal transcription apparatus

11. Post-Transcriptional Regulation: Alternative RNA Splicing

12. Post-Transcriptional Regulation: RNA Editing
Base substitution after transcription =

13. Translational Regulation: RNA Stability
Degradation of mRNA can occur from the 5’ or 3’ end
Stability of mRNA depends on
5’ cap
3’ poly-A tail
5’ and 3’ UTRs: serve as binding sites for regulatory factors
Coding region
Example: Hormone prolactin increases the longevity of casein mRNA coding for milk protein in lactating mammals

14. Translational Regulation
Masking of mRNAs
Many species store mRNAs in the cytoplasm of the egg. These mRNAs are inactive due to masking by proteins. Fertilization of the egg initiates unmasking and translation of these mRNAs.
Availability of specific tRNAs
In the embryonic development of a hornworm, an mRNA is present from day 1 but a specific tRNA needed for its translation is not produced until day 6.

15. Translational Regulation: RNA Silencing
Small interfering RNAs
microRNAs
RNA-induced silencing complex

16. Post-Translational Modification: Phosphorylation
Addition or removal of a phosphate group is a common way to change protein activity.

17. Post-Translational Modification: Peptide cleavage
Proteins that have an inactive form after synthesis are activated by removal of a small number of amino acids.
Prothrombin
Thrombin
Cleavage
Fibrinogen
Fibrin
Cleavage
Fibrin polymer (blood clot)
Activation of blood clotting factors by cleavage