620512FAd01 - A high-resolution map of active promoters in...

Info iconThis preview shows pages 1–2. Sign up to view the full content.

View Full Document Right Arrow Icon

Info iconThis preview has intentionally blurred sections. Sign up to view the full version.

View Full DocumentRight Arrow Icon
This is the end of the preview. Sign up to access the rest of the document.

Unformatted text preview: A high-resolution map of active promoters in the human genome Tae Hoon Kim 1,5 , Leah O. Barrera 1,5 , Ming Zheng 2 , Chunxu Qu 1 , Michael A. Singer 3 , Todd A. Richmond 3 , Yingnian Wu 2 , Roland D. Green 3 , and Bing Ren 1,4,6 1 Ludwig Institute for Cancer Research, 9500 Gilman Drive, La Jolla, CA 92093-0653, USA 2 8125 Math Sciences Building, UCLA Department of Statistics, Los Angeles, CA 90095-1554 3 Nimblegen Systems, Inc., 1 Science Court, Madison, WI 53711 4 Department of Cellular and Molecular Medicine, UCSD School of Medicine, 9500 Gilman Drive, La Jolla, CA 92093-0653, USA Abstract In eukaryotic cells, transcription of every protein-coding gene begins with the assembly of an RNA Polymerase II preinitiation complex (PIC) on the promoter 1 . The promoters, in conjunction with enhancers, silencers and insulators, define the combinatorial codes that specify gene expression patterns 2 . Our ability to analyze the control logic encoded in the human genome is currently limited by a lack of accurate information of the promoters for most genes 3 . Here, we describe a genome- wide map of active promoters in human fibroblast cells, determined by experimentally locating the sites of PIC binding throughout the human genome. This map defines 10,571 active promoters corresponding to 6,763 known genes and at least 1,199 un-annotated transcriptional units. Features of the map suggest extensive usage of multiple promoters by the human genes and widespread clustering of active promoters in the genome. In addition, examination of the genome-wide expression profile reveals four general classes of promoters that define the transcriptome of the cell. These results provide a global view of the functional relationship among the transcriptional machinery, chromatin structure and gene expression in human cells. The PIC consists of the RNA Polymerase II (RNAP), the transcription factor IID (TFIID) and other general transcription factors 4 . Our strategy to map the PIC binding sites involves a chromatin immunoprecipitation coupled DNA microarray analysis (ChIP-on-chip), which combines the immunoprecipitation of PIC-bound chromatin from formaldehyde crosslinked cells with parallel identification of the resulting bound DNA sequences using DNA microarrays 5,6 . Previously, we have demonstrated the feasibility of this strategy by successfully mapping active promoters in 1% of the human genome that correspond to the 44 genomic loci known as the ENCODE regions 6,7 . To apply this strategy to the entire human genome, we fabricated a series of DNA microarrays 8 containing roughly 14.5 million 50-mer oligonucleotides, designed to represent all the non-repeat DNA throughout the human genome at 100 basepairs (bp) resolution. We immunoprecipitated TFIID-bound DNA from the primary fibroblast IMR90 cells with a monoclonal antibody that specifically recognizes the TAF1 subunit of this complex (TBP associated factor 1, formerly TAF II 250 9, Fig 1a). We then amplified and fluorescently labeled the resulting DNA, and hybridized it to the above...
View Full Document

This note was uploaded on 05/28/2010 for the course WE BIBI010000 taught by Professor Marnikvuylsteke during the Spring '10 term at Ghent University.

Page1 / 10

620512FAd01 - A high-resolution map of active promoters in...

This preview shows document pages 1 - 2. Sign up to view the full document.

View Full Document Right Arrow Icon
Ask a homework question - tutors are online