METT-10, A Putative Methyltransferase, Inhibits Germ Cell Proliferative Fate in Caenorhabditis elegans

Germ-line stem cells are unique because they either self-renew through mitosis or, at a certain frequency, switch to meiosis and produce gametes. The switch from proliferation to meiosis is tightly regulated, and aberrations in switching result in either too little or too much proliferation. To understand the genetic basis of this regulation, we characterized loss-of-function mutations and a novel tumorous allele of Caenorhabditis elegans mett-10, which encodes a conserved putative methyltransferase. We show that METT-10 is a nuclear protein that acts in the germ line to inhibit the specification of germ-cell proliferative fate. METT-10 also promotes vulva, somatic gonad, and embryo development and ensures meiotic development of those germ cells that do differentiate. In addition, phenotypic analysis of a mett-10 null allele reveals that METT-10 enables mitotic cell cycle progression. The finding that METT-10 functions to inhibit germ-cell proliferative fate, despite promoting mitotic cell cycle progression of those germ cells that do proliferate, separates the specification of proliferative fate from its execution

High-Throughput Isolation of Caenorhabditis elegans Deletion Mutants

The nematode Caenorhabditis elegans is the first animal whose genome is completely sequenced, providing a rich source of gene information relevant to metazoan biology and human disease. This abundant sequence information permits a broad-based gene inactivation approach in C. elegans, in which chemically mutagenized nematode populations are screened by PCR for deletion mutations in a specific targeted gene. By handling mutagenized worm growth, genomic DNA templates, PCR screens, and mutant recovery all in 96-well microtiter plates, we have scaled up this approach to isolate deletion mutations in >100 genes to date. Four chemical mutagens, including ethyl methane sulfonate, ethlynitrosourea, diepoxyoctane, and ultraviolet-activated trimethylpsoralen, induced detectable deletions at comparable frequencies. The deletions averaged ∼1400 bp in size when using a ∼3 kb screening window. The vast majority of detected deletions removed portions of one or more exons, likely resulting in loss of gene function. This approach requires only the knowledge of a target gene sequence and a suitable mutagen, and thus provides a scalable systematic approach to gene inactivation for any organism that can be handled in high density arrays