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Sig (silencers in germline) mutants are defective in both germline transgene silencing and the maintenance of germ cell lineage. These mutants thus provide a viable genetic model for investigating germ cell biology, and possibly germ cell totipotency. We hypothesized that sig-2 affects germline function in C. elegans through a role in chromatin assembly. We therefore tested if sig-2 mutants exhibit defects in chromatin structure and organization compared to wild type animals. Transgenes are carried "extrachromosomally" as high-copy arrays in C. elegans. These extrachromosomal arrays are silenced in germ cells of wild type animals, but become desilenced in sig-2 mutants. Antibody analyses revealed that nucleosomal core histones on transgenes in normal germ cells are underacetylated relative to those on chromosomes. In contrast, transgene histones in sig-2 germ cells are detectably hyperacetylated. Interestingly, fluorescence light microscopy of DAPI stained sig-2 germ cells also indicates dysfunctional synapsis or pairing of meiotic chromosomes. These data suggest that normal SIG-2 function is required for some aspect of germ cell chromatin assembly that affects both meiotic synapsis and genetic silencing. These findings may shed light on a role for chromatin regulation in the maintenance of germ cell totipotency during development. Germ cells are the progenitors of all cell types in metazoan organisms. Germ cells remain totipotent throughout the lifetime of an organism; they maintain the ability to differentiate into any type of cell present in the organism, and are thus the ultimate type of stem cell. The mechanisms that allow germ cells to retain totipotency throughout development are poorly understood. Understanding fundamental characteristics that differ between germ cells and all somatic tissues may point to aspects of germ cell biology that affect totipotency. One obvious difference between the soma and germ cells in C. elegans is that transgene expression is strongly suppressed in the germ line, but not in the soma1. Mutants in this germ/soma distinction, called sig (silencers in germline) mutants, exhibit defects in germ cell function and viability, but no apparent somatic defects. This indicates that germ line silencing mechanisms are essential for germ cell function. What is the biological role of the SIG factors? One potential role is in chromatin organization. The activation of transgenes in the sig mutant germ cells indicates a change in chromatin configuration. One such change could be an alteration of chromatin protein modification, such as nucleosomal histone acetylation, which is known to correlate with expression competence. To investigate the role of sig genes in germ cell function, we have begun a characterization of one of the sig mutations, sig-2. Genetic and cytological investigations will be performed to examine the phenotypic consequences of the sig-2 mutation on germ cell function and chromatin organization.
A functional SIG-2 factor is required for germline silencing and maintaining normal fertility.
The sig-2 allele studied is recessive
SIG-2 is not maternally required
Transgene histones that are normally underacetylated become hyperacetylated in the sig-2 mutant
Homologous chromosome synapsis is impaired in sig-2 animals These results suggest that the normal function of SIG-2 is involved in aspects of chromatin assembly that affect both meiotic chromosome synapsis and germ cell silencing.
The authors would like to thank Kelvin Whisenhunt and Suzanne Roper for their unerring support and humor throughout this summer. The authors would also like to thank the Hughes Science Initiatives for providing the majority of the funding through the Howard Hughes Medical Institute, the National Science Foundation and the Graduate Division of Biological and Biomedical Sciences at Emory University.
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