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

2. http://cs273a.stanford.edu [Bejerano Aut08/09] 2 Lecture 8 Chains & Nets cont’d Conservation and Function Gene Regulation

3. Chromosome Mutations Five types exist: –Deletion –Inversion –Translocation –Nondisjunction –Duplication

4. 4 A Gene tree evolves with respect to a Species tree Species tree Gene tree Speciation Duplication Loss (deletion)

5. http://cs273a.stanford.edu [Bejerano Aut08/09] 5 Chains & Nets

6. http://cs273a.stanford.edu [Bejerano Aut08/09] 6 Net highlights rearrangements A large gap in the top level of the net is filled by an inversion containing two genes. Numerous smaller gaps are filled in by local duplications and processed pseudo-genes.

7. http://cs273a.stanford.edu [Bejerano Aut08/09] 7 Useful in finding pseudogenes Ensembl and Fgenesh++ automatic gene predictions confounded by numerous processed pseudogenes. Domain structure of resulting predicted protein must be interesting!

8. http://cs273a.stanford.edu [Bejerano Aut08/09] 8 And Retrogenes

9. http://cs273a.stanford.edu [Bejerano Aut08/09] 9 Conservation Track Documentation

10. http://cs273a.stanford.edu [Bejerano Aut08/09] 10 A Rearrangement Hot Spot Rearrangements are not evenly distributed. Roughly 5% of the genome is in hot spots of rearrangements such as this one. This 350,000 base region is between two very long chains on chromosome 7.

11. http://cs273a.stanford.edu [Bejerano Aut08/09] 11 Cautionary Note 1

12. http://cs273a.stanford.edu [Bejerano Aut08/09] 12 Cautionary Note 2

13. http://cs273a.stanford.edu [Bejerano Aut08/09] 13 Same Region… same in all the other fish

14. http://cs273a.stanford.edu [Bejerano Aut08/09] 14 Orthology vs. Paralogy

15. http://cs273a.stanford.edu [Bejerano Aut08/09] 15 Meet Your Genome contd. [Human Molecular Genetics, 3rd Edition]

16. http://cs273a.stanford.edu [Bejerano Aut08/09] 16 Sequence Conservation implies Function (but which function/s?...) human another species common ancestor...CTTTGCGA-TGAGTAGCATCTACTATTT......ACGTGGGACTGACTA-CATCGACTACGA... functional region! Comparative Genomics of Distantly related species: Note: the inverse “no conservation  no function” is a much weaker statement given current knowledge

17. http://cs273a.stanford.edu [Bejerano Aut08/09] 17 Our Place in the Tree of Life [Human Molecular Genetics, 3rd Edition]  you are here Which species to compare to? Too close and purifying selection will be largely indistinguishable from the neutral rate. Too far and many functional orthologs will diverge beyond our ability to accurately align them.

18. http://cs273a.stanford.edu [Bejerano Aut08/09] 18 Metazoans (multi-cellular organisms) [Human Molecular Genetics, 3rd Edition]  you are here

19. http://cs273a.stanford.edu [Bejerano Aut08/09] 19 Vertebrates: what to sequence? [Human Molecular Genetics, 3rd Edition]  you are here, Opossum, Lizard, Stickleback too far sweet spot too close

20. http://cs273a.stanford.edu [Bejerano Aut08/09] 20 The Dawn of Whole Genome Comparative Genomics 2001 2002 40% DNA alignable 95% coding genes shared

21. http://cs273a.stanford.edu [Bejerano Aut08/09] 21 More Species Have Joined Since

22. http://cs273a.stanford.edu [Bejerano Aut08/09] 22 How They Measured all human-mouse alignments human-mouse ancestral repeats alignment Difference: 5% of Human Genome [Mouse consortium, Nature 2002]

23. http://cs273a.stanford.edu [Bejerano Aut08/09] 23 Conserved elements in the Human Genome all human-mouse alignments human-mouse ancestral repeats alignment Difference: 5% of Human Genome [Mouse consortium, Nature 2002] election human-mouse ancestral repeats alignment 85%id on average Ultraconservation

24. http://cs273a.stanford.edu [Bejerano Aut08/09] 24 Ultraconserved Elements [Bejerano et al., Science 2004] fish 481 elements perfectly conserved (100%id) over 200bp or more between human, mouse and rat.

25. http://cs273a.stanford.edu [Bejerano Aut08/09] 25 Ultraconserved Elements: Why? Hundreds of long substrings identical between amniotes  they must have rejected many different changes. But... all functions we understand in our genome are encoded using redundant codes. E.g. Protein Coding Genes: DNA – 10 8 letters over alphabet of 4. Protein – 10 2 letters over alphabet of 20. Coding: 3 DNA letters → 1 Protein letter. * * * * * [Bejerano et al., Science 2004]

26. http://cs273a.stanford.edu [Bejerano Aut08/09] 26 No known function requires this much conservation CDSncRNATFBS * * * * * seq. ?

27. http://cs273a.stanford.edu [Bejerano Aut08/09] 27 Other Ways to Measure [Lunter et al, 2006] [Cooper et al., 2005]

28. http://cs273a.stanford.edu [Bejerano Aut08/09] 28, Opossum, Lizard, Stickleback Phylogenetic Shadowing [Human Molecular Genetics, 3rd Edition]  you are here too close “too close” can actually be a boon if you have enough closely related genomes

29. http://cs273a.stanford.edu [Bejerano Aut08/09] 29 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?

30. http://cs273a.stanford.edu [Bejerano Aut08/09] 30 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

31. http://cs273a.stanford.edu [Bejerano Aut08/09] 31 Gene regulation = when/where to make protein gene (how to) control region (when & where) DNA ~10 3 letters

32. http://cs273a.stanford.edu [Bejerano Aut08/09] 32 Vertebrate Gene Regulation gene (how to) control region (when & where) DNA proximal: in 10 3 letters distal: in 10 6 letters DNA binding proteins

33. http://cs273a.stanford.edu [Bejerano Aut08/09] 33 unicellular multicellular Unicellular vs. Multicellular

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

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

36. http://cs273a.stanford.edu [Bejerano Aut08/09] 36 Enhancer verification Matched staining in genital eminence Matched staining in dorsal apical ectodermal ridge (part of limb bud)