Regulatory Elements and Gene Expression in Primates and Diverse Human Cell-types

Abstract

After finishing a human genome reference sequence in 2002, the genomics community hasturned to the task of interpreting it. A primary focus is to identify and characterize not onlyprotein-coding genes, but all functional elements in the genome. The effort has identifiedmillions of regulatory elements across species and in hundreds of human cell-types. Nearlyall identified regulatory elements are found in non-coding DNA, hypothesizing a functionfor previously unannotated sequence. The ability to identify regulatory DNA genome-wideprovides a new opportunity to understand gene regulation and to ask fundamental questionsin diverse areas of biology.One such area is the aim to understand the molecular basis for phenotypic differencesbetween humans and other primates. These phenotypic differences are partially drivenby mutations in non-coding regulatory DNA that alter gene expression. This hypothesishas been supported by differential gene expression analyses in general, but we have notyet identified specific regulatory variants responsible for differences in transcription andphenotype. I have worked to identify regulatory differences in the same cell-type isolatedfrom human, chimpanzee, and macaque. Most regulatory elements were conserved amongall three species, as expected based on their central role in regulating transcription. How-ever, several hundred regulatory elements were gained or lost on the lineages leading tomodern human and chimpanzee. Species-specific regulatory elements are enriched neardifferentially expressed genes, are positively correlated with increased transcription, showevidence of branch-specific positive selection, and overlap with active chromatin marks.ivSpecies-specific sequence differences in transcription factor motifs found within this regu-latory DNA are linked with species-specific changes in chromatin accessibility. Together,these indicate that species-specific regulatory elements contribute to transcriptional andphenotypic differences among primate species.Another fundamental function of regulatory elements is to define different cell-types inmulticellular organisms. Regulatory elements recruit transcription factors that modulategene expression distinctly across cell-types. In a study of 112 human cell-types, I classifiedregulatory elements into clusters based on regulatory signal tissue specificity. I then usedthese to uncover distinct associations between regulatory elements and promoters, CpG-islands, conserved elements, and transcription factor motif enrichment. Motif analysisidentified known and novel transcription factor binding motifs in cell-type-specific andubiquitous regulatory elements. I also developed a classifier that accurately predicts cell-type lineage based on only 43 regulatory elements and evaluated the tissue of origin forcancer cell-types. By correlating regulatory signal and gene expression, I predicted targetgenes for more than 500k regulatory elements. Finally, I introduced a web resource toenable researchers to explore these regulatory patterns and better understand how expressionis modulated within and across human cell-types.Regulation of gene expression is fundamental to life. This dissertation uses identifiedregulatory DNA to better understand regulatory systems. In the context of either evolution-ary or developmental biology, understanding how differences in regulatory DNA contributeto phenotype will be central to completely understanding human biology.