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

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

3. http://cs273a.stanford.edu [Bejerano Aut08/09] 3 Vertebrate Gene Regulation gene (how to) control region (when & where) DNA proximal: in 10 3 letters distal: in 10 6 letters DNA binding proteins

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

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

6. 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

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

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

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

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

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

12. 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] 13

14. 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)

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

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

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

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

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

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

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

22. 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]

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

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

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

26. http://cs273a.stanford.edu [Bejerano Aut08/09] 26 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] Verifying repressors is trickier [loss vs. gain of function]. How do you predict an enhancer from a repressor? Duh... repressors Repressors

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

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

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

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

31. http://cs273a.stanford.edu [Bejerano Aut08/09] 31 Human Genome: 3*10 9 letters What They Found [Science 2004 Breakthrough of the Year, 5 th runner up] 1.5% known function >50% junk 3x more functional DNA than known! compare to other species >5% human genome functional ~10 6 substrings do not code for protein What do they do then?

32. http://cs273a.stanford.edu [Bejerano Aut08/09] 32 Gene number does not correlate with Complexity Gene families are important. Many are surprisingly old. But - fly worm human weed fish rice # genes 10 3 cells 10 13 cells pre-genomic era: “100,000 genes to the human genome” last count down to 20,500 human genes

33. http://cs273a.stanford.edu [Bejerano Aut08/09] 33 Most Non-Coding Elements are likely cis-regulatory 9Mb “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

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

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

36. 36 Cell Communication Lodish, 20-1

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

38. 38 Wnt and Hedgehog signaling Jacob & Lum Science 2007