Identification of DNA Elements Governing Chromatin Function in C. elegans

Jason Lieb (PI)	University of North Carolina, Chapel Hill
Julie Ahringer	University of Cambridge
Abby Dernburg	University of California, Berkeley
Arshad Desai	University of California, San Diego
Roland Green	NimbleGen
Xiaole Liu	Dana Farber Cancer Institute
Eran Segal	Weizmann Institute
Susan Strome	University of California, Santa Cruz

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Eukaryotic genomes are packaged into chromatin, which regulates the function of proteins that mediate transcriptional activity and other essential processes, including recombination and the faithful segregation of the genome during mitosis and meiosis. The goal of this proposal is to identify discrete elements that regulate chromatin structure and function in the nematode C. elegans, a model metazoan of central importance in large-scale genomic research and gene function discovery. We will first use ChIP-chip and related methods to map the genomic distributions of selected histone modifications and chromosome-associated proteins, and then use that information, in combination with data from other modENCODE groups, to build quantitative models of chromatin function. Specifically, we will:

1. Identify and technically validate functional elements that control chromatin and chromosome behavior.
   The focus of our analysis will be elements that specify nucleosome positioning and occupancy, control domains of gene expression, induce repression of the X chromosome, guide mitotic segregation and genome duplication, govern homolog pairing and recombination during meiosis, and organize chromosome positioning within the nucleus. 126 strategically selected targets include key histone modifications, histone variants, RNA polymerase II isoforms, dosage-compensation proteins, centromere components, homolog-pairing facilitators, recombination markers, and nuclear-envelope constituents. An efficient pipeline design will facilitate identification and validation of the different classes of functional elements associated with these targets and integrate the results with the well-annotated C. elegans genome.
2. Biologically validate identified functional elements and build integrated, quantitative models of chromo-some function.
   We will integrate information generated in Aim 1 with existing knowledge on the biology of the targets, perform ChIP-chip analysis on mutant and RNAi extracts lacking selected target proteins, use extrachromosomal arrays to assess the ability of candidate identified sequence motifs to recruit targets in vivo, identify tissue-specific patterns of selected targets, and create integrated, quantitative models of transcription and whole-chromosome functions.

Achieving these goals in the context of the ongoing expansion and rich history of C. elegans research will provide an important milestone in meeting the challenge of using genome sequence information to understand and predict biological functions.

Experimental Approaches

1. ChIP-Chip to map genomic locations of modified histones and chromosome-associated proteins, and to assess changes induced by mutations and RNAi knockdowns of selected target genes.
2. Extrachromosomal arrays to assess ability of candidate sequence motifs to recruit target proteins in vivo and to identify tissue-specific expression patterns of targets.

Information Resources to be Generated

• ChIP-chip data on Nimblegen tiling arrays (2.1 M probes, ~50 bp resolution) which is amenable to track-type display.
• Nucleosome mapping data done at higher resolution (10.5 M probes, ~10 bp resolution).
• Analysis tracks, including peak calls or other annotations.
• RNA expression data from promoters fused to reporter genes and expressed in extrachromosomal arrays.

Contact Information

• Jason Lieb (PI, C. elegans ChIP-chip, Somatic Dosage Compensation and Chromatin) 919-843-3228 (office)
• Julie Ahringer (Chromatin Regulation and Development) +44 1223 334088
• Abby Dernburg (meiotic pairing and recombination) 510-495-2306 (office)
• Arshad Desai (kinetochore proteins; centromeres) 858-552-4920 x7821 (office) 858-552-4920 x7890 (lab)
• Xiaole Liu (ChIP-chip data analysis and DNA sequence motif finding (617) 632-2472 (office)
• Eran Segal (computational models of chromatin and transcriptional control) +972-8-934-4282 (office) +7 hours East Coast time +10 hours West Coast time
• Susan Strome (chromatin and gene regulation in meiosis)831-459-5683 (office)
• Roland Green and Xinmin Zhang
• Ercan Sevinc (Informatics)