Profs: Serafim Batzoglou, Gill Bejerano TAs: Cory McLean, Aaron Wenger 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]

Lecture 18 Guest Talks Ultraconservation The Projects http://cs273a.stanford.edu [Bejerano Aut08/09]

Comparative Genomics what made us human? * t mouse rat chimp chicken fugu zfish dog tetra opossum cow macaque platypus * While we may not agree about everything… Incomplete list: also cat, armadillo, elephant, rabbit, shrew, tenrec, wallaby (2x) Some not mammals, obviously, but useful outgroups in phylogenetic analysis of mammals t http://cs273a.stanford.edu [Bejerano Aut08/09]

Genotype Phenotype http://cs273a.stanford.edu [Bejerano Aut08/09] With the vast genomic resources available nowadays, we can isolate both genotypic and phenotypic differences between species. However, a gap still exists in our ability to associate genotype with phenotype, and an overarching goal in biology today is to start bridging that gap, to be able to say: This inversion causes foot shape alterations This duplication and that substitution both play a role in brain size Or going from phenotype to genotype, This sweat-related gene has some change in chimpanzee relative to human The study I am talking about today tries to link genotype to phenotype for a single type of genomic rearrangement, human-specific deletions. Genotype Phenotype 4 http://cs273a.stanford.edu [Bejerano Aut08/09] 4

Comparative Genomics How did some of our relatives go back? * t human mouse rat chimp chicken fugu zfish dog tetra opossum cow macaque platypus * While we may not agree about everything… Incomplete list: also cat, armadillo, elephant, rabbit, shrew, tenrec, wallaby (2x) Some not mammals, obviously, but useful outgroups in phylogenetic analysis of mammals t http://cs273a.stanford.edu [Bejerano Aut08/09]

Now at the Species Level How did we leave the ocean? human mouse rat chimp chicken fugu zfish dog tetra opossum cow macaque platypus * While we may not agree about everything… Incomplete list: also cat, armadillo, elephant, rabbit, shrew, tenrec, wallaby (2x) Some not mammals, obviously, but useful outgroups in phylogenetic analysis of mammals t http://cs273a.stanford.edu [Bejerano Aut08/09]

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

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

Typical DNA Conservation levels Conserved elements between human and mouse are on average 85% identical. [mouse consortium, 2002] http://cs273a.stanford.edu [Bejerano Aut08/09] [Bejerano et al., Science 2004]

Ultraconserved Elements fish http://cs273a.stanford.edu [Bejerano Aut08/09] [Bejerano et al., Science 2004]

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

Discovery can be fun http://cs273a.stanford.edu [Bejerano Aut08/09] (compared to 4 results day before our ScienceExpress paper) 31,800 http://cs273a.stanford.edu [Bejerano Aut08/09]

Predictions and Proofs I Based on public domain genome wide data: ultraconserved elements one subset codes protein larger subset does not generate testable hypotheses for function from existing knowledge (2004) [Pennacchio et al., Nature, 2006] http://cs273a.stanford.edu [Bejerano Aut08/09]

Predictions and Proofs II Based on public domain genome wide data: ultraconserved elements one subset codes protein larger subset does not generate testable hypotheses for function from existing knowledge (2004) post transcriptional modification [Pennacchio et al., Nature, 2006] http://cs273a.stanford.edu [Bejerano Aut08/09]

Of the 29 exonic ultraconserved elements in RNA-binding protein genes in human, 15 have human and/or mouse EST evidence suggesting the presence of AS-NMD in those regions. [Ni et al., Genes & Dev 2007 ] http://cs273a.stanford.edu [Bejerano Aut08/09]

Model for Homeostatic Auto/Cross-regulation [Ni et al., Genes & Dev 2007 ] http://cs273a.stanford.edu [Bejerano Aut08/09]

Ultras are Under Strong Human Selection Mutational cold spots? NO. Rare (new) mutations are introduced to the population. Fierce purifying selection? YES. Very few of these get anywhere near fixation. A A G A A humans chimp Ultra DAF NonSyn DAF [Katzman et al, Science ,2007] http://cs273a.stanford.edu [Bejerano Aut08/09]

Genomic Distribution of Ultraconserved Elements exonic non possibly Origins? http://cs273a.stanford.edu [Bejerano Aut08/09]

UC.338 comes from an ancient repeat ultraconserved exon novel coelacanth repeat enhancer LF-SINE [Bejerano et al, Nature ,2006] http://cs273a.stanford.edu [Bejerano Aut08/09]

Touch an Ultra And You - DIY No Apparent Phenotype 4-8% of documented mouse KO genes display no phenotype (inc. Pbx2, Nkx6.2, Gli1) http://cs273a.stanford.edu [Bejerano Aut08/09] [Ahituv et al., PLoS Biology, 2007]

Hypothesis Can ultraconserved element mutations be under extreme purifying selection but the whole element be expendable? http://cs273a.stanford.edu [Bejerano Aut08/09]

What did Mother Nature favor? DNA element t mouse http://cs273a.stanford.edu [Bejerano Aut08/09]

Quite Some Time Later http://cs273a.stanford.edu [Bejerano Aut08/09] The horror … http://cs273a.stanford.edu [Bejerano Aut08/09]

Ultras are Fiercely Retained No Apparent Phenotype rodents deleted 11/15157 = 0.072% Ultras are >300 fold more persistent than neutral DNA But Doomed ... (25% deleted) http://cs273a.stanford.edu [Bejerano Aut08/09]

Enough about us…. http://cs273a.stanford.edu [Bejerano Aut08/09]

What we do understand.. Ultraconserved elements exist. They are maintained via strong on-going selection. It is a heterogeneous bunch: Some mediate splicing Some regulate gene expression Some express ncRNAs (categories are not necessarily mutually exclusive) Knockouts of four regulatory ultras did not lead to severe phenotypes (similar protein cases: Pbx2, Nkx6.2, Gli1) http://cs273a.stanford.edu [Bejerano Aut08/09]

What we don’t understand Their functional density: How did they come to be? What is the selective advantage that lets them persist? http://cs273a.stanford.edu [Bejerano Aut08/09]

Broad Guess It’s about 3-D structure. Observation: rDNA (18S, 28S) have ultraconserved stretches, multiple constraints in a complex 3-D structure, the Ribosome. ncRNA ultras: structure confers function Splicing related ultras: the Splicosome Cis-reg ultras: TSS 3-D proximity, chromatin and/or packed TFBS (Transcription factories?) TSS http://cs273a.stanford.edu [Bejerano Aut08/09]