ANALYSIS OF CONTEXT-DEPENDENT MODULATION OF EPIGENETIC GENE ACTIVITIES IN CAENORHABDITIS ELEGANS
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Date
2007-01-25T21:06:23Z
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Johns Hopkins University
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.
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Keywords
Epigenetics, Gene silencing, Chromatin, C elegans, Transgenes, Imprinting