Control of eukaryotic gene expression

1. Control of eukaryotic gene expression • As usual, much more complicated than in prokaryotes. • Increased amount of DNA • Tight packing into nucleosomes • Physical separation between nucleus & ribosomes • mRNA processing and different life span • Multicellular w/ development & differentiation • Many options for control • From DNA rearrangements to protein modification

2. Overview of critical elements • DNA regions • Promoter: TATA, CAAT, & GCG boxes • RNA polymerase binds, with assistance. • Enhancers and silencers: short sequences that proteins bind to that influence transcription. • Proteins • TFIID: made of TBP (TATA box binding protein) and about 12 TAFs (TBP associated factors). • Transcription factors: basal and enhancer binding • RNA polymerase II (for mRNA)

3. The Process of Transcription • Promoter recognition: 2 consensus sequences • The -10 region: TATAAT (10 bases upstream from where transcription actually starts. • The -35 region, farther upstream, also important. • “Consensus” sequence meaning the DNA sequence from many genes averages out to this. • The closer these 2 regions actually are to the consensus sequences, the “stronger” the promoter, meaning the more likely RNA polymerase binding and transcription will occur.

4. Consensus sequence Numbers indicate the percentage of different genes in which that nucleotide appears in that spot in the promoter sequence. http://www.uark.edu/campus-resources/mivey/m4233/promoter.gif

5. DNA regions- eukaryotes Binding of factors to the TATA box area essential for transcription to occur. Binding of factors to the promoter influence how much transcription occurs. http://web.indstate.edu/thcme/mwking/gene-regulation.html

6. Promoter and protein factors TATA box CAAT box Other promoter elements such as hormone response elements.

7. What happens at the promoter • TFIID: a multicomponent protein made of • TBP: TATA Box Binding Protein • Actually binds to the TATA box on DNA • TAFs: TBP Associated Factors • Ten to 14 proteins that bind to the TBP • This complex , along with other TFs, recruits the RNA pol II so it can begin transcription • Unlike on prokaryotes, the RNA Pol II does not actually bind to the TATA box of promoter. See also cats.med.uvm.edu/.../ 2.1.grg.promoter.html

8. What happens at promoter Based on: http://www.nig.ac.jp/section/mitsuzawa/mitsuzawa_fig.jpg

9. Enhancers • Short DNA segments that enhance transcription • “silencers” apparently exert negative control • Important aspects of enhancers • Specificity: certain sequences bind to certain transcription factors • Can be located upstream, downstream, within the gene, at a considerable distance from promoter. • Can be inverted without changing affect. • Promote looping of DNA, a 3D change that somehow promotes transcription.

10. How enhancers work Specificity: Depending on why gene is needed in the cell, a unique enhancer sequence is bound to by one transcription factor, but not another. Multiple enhancers: transcription can be increased by several different signals (transcription factors binding to several enhancers). http://www.emunix.emich.edu/~rwinning/genetics/eureg3.htm

11. How enhancers work-2 Enhancer promotes DNA looping which leads to increased transcription. The “enhancer-binding protein” shown would also be called a transcription factor in most textbooks. http://users.rcn.com/jkimball.ma.ultranet/BiologyPages/P/Promoter.html#enhancers

12. More on Transcription Factors • Proteins that bind to DNA • Helix-turn-helix • Zincfinger • Leucine zipper and other unique-named motifs • TFs typically have more than 1 domain, also bind to other proteins. • Bind at core promoter, at upstream promoter elements, or at enhancers. http://www.web-books.com/MoBio/Free/images/Ch4F5b.gif

13. For more viewing • www.bmb.psu.edu/.../ tan/lab/gallery_protdna.html • 3D imaging of transcription factor binding to DNA. Leucine zipper http://www.uic.edu/classes/bios/bios100/mike/spring2003/leucinezipper.jpg

14. Other controlling factors • Chromatin remodeling • Chromatin remodeling complexes • Specific proteins disrupt nucleosomes, make DNA available to be transcribed • Acetylation of histones • Causes histones to bind DNA less tightly • Methylation: nearly universal way of decreasing use of DNA • Methyl group blocks access of proteins • Barr body, other un-transcribed DNA highly methylated.

15. Two short examples • Endocrine tissues send a chemical signal to a target tissue, how does this effect gene expression? • Steroid hormone • Lipid substance, diffuses through cell membrane • Binds to its receptor = a transcription factor. • Enters nucleus, turns on appropriate genes. • Binds to enhancers/promoter elements • Peptide hormone • Binds to receptor on cell surface • Triggers 2nd messenger which activates a transcription factor.

16. Other methods of genetic control • DNA changes • rearrangements (e.g. antibody genes) • Methylation of C (many GC rich regions) • At level of mRNA • Differential splicing: different proteins. • De-adenylate tail, decap: nuclease destroy mRNA. • At the level of Translation • mRNA can be sequestered, used later. • Lot of mRNA means lots of translation, product • Post-translational modification • Phosphorylation, methylation, acetylation, etc.