ANALYSIS OF CONTEXT-DEPENDENT MODULATION OF EPIGENETIC GENE ACTIVITIES IN CAENORHABDITIS ELEGANS

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Title: ANALYSIS OF CONTEXT-DEPENDENT MODULATION OF EPIGENETIC GENE ACTIVITIES IN CAENORHABDITIS ELEGANS
Author: Sha, Kyle
Abstract: We initiated a screen for trans-acting factors that modulate expression of a transgene reporter in C. elegans. In the process, we fortuitously generated transgene derivatives that exhibited an imprinting effect. Imprinting is the non-equivalence of reciprocal matings. From a single progenitor line carrying an extra-chromosomal unc-54::gfp transgene array, we generated three independent autosomal integrations of the unc-54::gfp transgene. The progenitor line, two of its three integrated derivatives, and a non-related unc-119::gfp fusion exhibit an imprinting effect: single-generation transmission of these transgenes through the male germline results in approximately 1.5-2.0 fold greater expression than transmission through the female germline. There is a detectable resetting of the imprint after passage through the opposite germline for a single generation, indicating that the imprinted status of the transgenes is reversible. In cases where the transgene is maintained in either the oocyte lineage or sperm lineage for multiple, consecutive generations, a full reset requires passage through the opposite germline for several generations. Taken together, our results indicate that C. elegans has the ability to imprint chromosomes and that differences in the cell and/or molecular biology of oogenesis and spermatogenesis are manifest in an imprint that can persist in both somatic and germline gene expression for multiple generations. To gain insight into C. elegans chromatin, we expressed the E. coli dam methyltransferase in C. elegans muscle cells in order to map susceptible regions of the genome to DNA modification. Dam methyltransferase catalyzes the transfer of a methyl from S-adenosyl-methionine to adenine in the target sequence GATC. Using restriction enzymes that are sensitive to the methylation status of GATC, we mapped 309 dam – iii – methyltransferase sites out of a potential total of 269,000 sites distributed more or less uniformly throughout the C. elegans genome. Our preliminary analysis indicated that targets of DAM are distributed uniformly throughout the entire genome, without any apparent bias towards specific chromosomes or genomic regions. SAGE analysis revealed that dam methyltransferase does not have any bias towards muscle-specific genes. Thus, our experimental system is a potentially useful tool for investigating genome-wide chromatin accessibility from within muscle tissue.
URI: http://jhir.library.jhu.edu/handle/1774.2/1337
Date: 2007-01-25
Subject: Epigenetics
Gene silencing
Chromatin
C elegans
Transgenes
Imprinting

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