Gene Regulation and Transcription Factors in Vertebrates

1. MW  11:00-12:15 in Beckman B302 Profs: Serafim Batzoglou, Gill Bejerano TAs: Cory McLean, Aaron Wenger http://cs273a.stanford.edu [Bejerano Aut08/09]

2. Lecture 12 • Gene Cis Regulation • Signal Transduction http://cs273a.stanford.edu [Bejerano Aut08/09]

3. Vertebrate Gene Regulation • gene (how to) • control region(when & where) distal: in 106 letters DNA DNA binding proteins proximal: in 103 letters http://cs273a.stanford.edu [Bejerano Aut08/09]

4. Unicellular vs. Multicellular unicellular multicellular http://cs273a.stanford.edu [Bejerano Aut08/09]

5. Vertebrate Transcription Regulation http://cs273a.stanford.edu [Bejerano Aut08/09]

6. Pol II Transcription • Key components: • Proteins • DNA sequence • DNA epigenetics • Protein components: • General Transcription factors • Activators • Co-activators http://cs273a.stanford.edu [Bejerano Aut08/09]

7. Activators & Co-Activators Protein - Protein Protein - DNA http://cs273a.stanford.edu [Bejerano Aut08/09]

8. The Core Promoter http://cs273a.stanford.edu [Bejerano Aut08/09]

9. Tx Factors Binding Sites http://cs273a.stanford.edu [Bejerano Aut08/09]

10. CpG islands http://cs273a.stanford.edu [Bejerano Aut08/09]

11. Chromatin Remodeling “off” “on” http://cs273a.stanford.edu [Bejerano Aut08/09]

12. Nucleosome tail modifications • Lysine acetylations. • Histone Acetyl-Transferases (HAT) & Histone Deacetylases (HDAC). • Lysine and Argenine Metylations. • Modified by histone-metyl-transferase. • Phosphorilation. • Ubiquitination. • H2A ubiquitination affects 10-15% of this histone in most eukaryotic cells • ADP-ribosylation.

13. http://cs273a.stanford.edu [Bejerano Aut08/09]

14. Cis-Regulatory Components • Low level (“atoms”): • Promoter motifs (TATA box, etc) • Transcription factor binding sites (TFBS) • Mid Level: • Promoter • Enhancers • Repressors/Silencers • Insulators/boundary elements • Cis-Regulatory Modules (CRM) • Locus Control Regions (LCR) • High Level: • Gene Expression Domains • Gene Regulatory Networks (GRN) http://cs273a.stanford.edu [Bejerano Aut08/09]

15. Distal Transcription Regulatory Elements http://cs273a.stanford.edu [Bejerano Aut08/09]

16. Enhancers http://cs273a.stanford.edu [Bejerano Aut08/09]

17. Enhancers: action over very large distances RNAP II Basal factors promoter Enhancer with bound protein http://cs273a.stanford.edu [Bejerano Aut08/09]

18. Transient Transgenic Enhancer Assay in situ Conserved Element Minimal Promoter Reporter Gene Construct is injected into 1 cell embryos Taken out at embryonic day 10.5-14.5 Assayed for reporter gene activity transgenic http://cs273a.stanford.edu [Bejerano Aut08/09]

19. Enhancer verification Matched staining in dorsal apical ectodermal ridge (part of limb bud) Matched staining in genital eminence http://cs273a.stanford.edu [Bejerano Aut08/09]

20. Vertebrate Enhancer Combinatorics brain limb neural tube Sall1 http://cs273a.stanford.edu [Bejerano Aut08/09]

21. Vertebrate Enhancer Combinatorics http://cs273a.stanford.edu [Bejerano Aut08/09]

22. What are Enhancers? • What do enhancers encode? • Surely a cluster of TF binding sites. • [but TFBS prediction is hard, fraught with false positives] • What else? DNA Structure related properties? • So how do we recognize enhancers? • Sequence conservation across multiple species • [weak but generic] http://cs273a.stanford.edu [Bejerano Aut08/09]

23. Gene Expression Domains: Independent http://cs273a.stanford.edu [Bejerano Aut08/09]

24. Gene Expression Domains: Dependent http://cs273a.stanford.edu [Bejerano Aut08/09]

25. Repressors / Silencers http://cs273a.stanford.edu [Bejerano Aut08/09]

26. What are Enhancers? Repressors • What do enhancers encode? • Surely a cluster of TF binding sites. • [but TFBS prediction is hard, fraught with false positives] • What else? DNA Structure related properties? • So how do we recognize enhancers? • Sequence conservation across multiple species • [weak but generic] • Verifying repressors is trickier [loss vs. gain of function]. • How do you predict an enhancer from a repressor? Duh... repressors repressors http://cs273a.stanford.edu [Bejerano Aut08/09]

27. Insulators http://cs273a.stanford.edu [Bejerano Aut08/09]

28. Transcription Regulation & Human Disease [Wang et al, 2000] http://cs273a.stanford.edu [Bejerano Aut08/09]

29. Other Positional Effects [de Kok et al, 1996] http://cs273a.stanford.edu [Bejerano Aut08/09]

30. TFs in the Human Genome http://cs273a.stanford.edu [Bejerano Aut08/09]

31. What They Found Human Genome: 3*109 letters 1.5% known function compare to other species >50% junk >5% human genome functional 3x more functional DNA than known! ~106 substrings do not code for protein What do they do then? [Science 2004 Breakthrough of the Year, 5th runner up] http://cs273a.stanford.edu [Bejerano Aut08/09]

32. Gene number does not correlate with Complexity Gene families are important. Many are surprisingly old. But - 1013 cells pre-genomic era: “100,000 genes to the human genome” 103 cells fly worm human weed fish rice last count down to 20,500 human genes # genes http://cs273a.stanford.edu [Bejerano Aut08/09]

33. Most Non-Coding Elements are likely cis-regulatory “IRX1 is a member of the Iroquois homeobox gene family. Members of this family appear to play multiple roles during pattern formation of vertebrate embryos.” gene deserts regulatory jungles 9Mb http://cs273a.stanford.edu [Bejerano Aut08/09]

34. Rapid TFBS turnover http://cs273a.stanford.edu [Bejerano Aut08/09]

35. Signal Transduction http://cs273a.stanford.edu [Bejerano Aut08/09]

36. Cell Communication Lodish, 20-1

37. Signaling Pathways Important in Developmental Biology • Wnt/Frizzled through b-catenin • Hedgehog • TGF-b family through Smads • Growth factors via JAK-STATs • Notch • Integrin • TNF

38. Wnt and Hedgehog signaling Jacob & Lum Science 2007