SUMMARYThe C. elegans germline provides an excellent model for analyzing the regulation of stem cell activity and the decision to differentiate and undergo meiotic development. The distal end of the adult hermaphrodite germline contains the proliferative zone, which includes a population of mitotically cycling cells and cells in meiotic S phase, followed by entry into meiotic prophase. The proliferative fate is specified by somatic distal tip cell (DTC) niche-germline GLP-1 Notch signaling through repression of the redundant GLD-1 and GLD-2 pathways that promote entry into meiosis. Here, we describe characteristics of the proliferative zone, including cell cycle kinetics and population dynamics, as well as the role of specific cell cycle factors in both cell cycle progression and the decision between the proliferative and meiotic cell fate. Mitotic cell cycle progression occurs rapidly, continuously, with little or no time spent in G1, and with cyclin E (CYE-1) levels and activity high throughout the cell cycle. In addition to driving mitotic cell cycle progression, CYE-1 and CDK-2 also play an important role in proliferative fate specification. Genetic analysis indicates that CYE-1/CDK-2 promotes the proliferative fate downstream or in parallel to the GLD-1 and GLD-2 pathways, and is important under conditions of reduced GLP-1 signaling, possibly corresponding to mitotically cycling proliferative zone cells that are displaced from the DTC niche. Furthermore, we find that GLP-1 signaling regulates a third pathway, in addition to the GLD-1 and GLD-2 pathways and also independent of CYE-1/CDK-2, to promote the proliferative fate/inhibit meiotic entry.
p53 is a tumor suppressor gene whose regulation is crucial to maintaining genome stability and for the apoptotic elimination of abnormal, potentially cancer-predisposing cells. C. elegans contains a primordial p53 gene, cep-1, that acts as a transcription factor necessary for DNA damage-induced apoptosis. In a genetic screen for negative regulators of CEP-1, we identified a mutation in GLD-1, a translational repressor implicated in multiple C. elegans germ cell fate decisions and related to mammalian Quaking proteins. CEP-1-dependent transcription of proapoptotic genes is upregulated in the gld-1(op236) mutant and an elevation of p53-mediated germ cell apoptosis in response to DNA damage is observed. Further, we demonstrate that GLD-1 mediates its repressive effect by directly binding to the 3'UTR of cep-1/p53 mRNA and repressing its translation. This study reveals that the regulation of cep-1/p53 translation influences DNA damage-induced apoptosis and demonstrates the physiological importance of this mechanism.
In C. elegans, the decision between germline stem cell proliferation and entry into meiosis is controlled by GLP-1 Notch signaling, which promotes proliferation through repression of the redundant GLD-1 and GLD-2 pathways that direct meiotic entry. We identify prp-17 as another gene functioning downstream of GLP-1 signaling that promotes meiotic entry, largely by acting on the GLD-1 pathway, and which also functions in female germline sex determination. PRP-17 is orthologous to the yeast and human pre-mRNA splicing factor PRP17/CDC40 and can rescue the temperature-sensitive lethality of yeast PRP17. This link to splicing led to an RNAi screen of predicted C. elegans splicing factors in sensitized genetic backgrounds. We found that many genes throughout the splicing cascade function in the proliferation/meiotic entry decision and germline sex determination indicating that splicing per se, rather than a novel function of a subset of splicing factors, is necessary for these processes.
Caenorhabditis elegans is an ideal organism for the study of the molecular basis of fundamental biological processes such as germline development, especially because of availability of the whole genome sequence and applicability of the RNA interference (RNAi) technique. To identify genes involved in germ-line development, we produced subtracted cDNA pools either enriched for or deprived of the cDNAs from germ-line tissues. We then performed differential hybridization on the high-density cDNA grid, on which about 7,600 nonoverlapping expressed sequence tag (EST) clones were spotted, to identify a set of genes specifically expressed in the germ line. One hundred and sixty-eight clones were then tested with the RNAi technique. Of these, 15 clones showed sterility with a variety of defects in germ-line development. Seven of them led to the production of unfertilized eggs, because of defects in spermatogenesis (4 clones), or defects in the oocytes (3 clones). The other 8 clones led to failure of oogenesis. These failures were caused by germ-line proliferation defect (Glp phenotype), meiotic arrest, and defects in sperm-oocyte switch (Mog phenotype) among others. These results demonstrate the efficacy of the screening strategy using the EST library combined with the RNAi technique in C. elegans.
Eukaryotic initiation factor 5A (eIF-5A) was originally isolated as a translation initiation factor. However, this function has since been reconsidered, with recent studies pointing to roles for eIF-5A in mRNA metabolism and trafficking [Microbiol. Mol. Biol. Rev. 66 (2002) 460; Eur. Mol. Biol. Org. J. 17 (1998) 2914]. The Caenorhabditis elegans genome contains two eIF-5A homologues, iff-1 and iff-2, whose functions in vivo were examined in this study. The iff-2 mutation causes somatic defects that include slow larval growth and disorganized somatic gonadal structures in hermaphrodites. iff-2 males show disorganized tail sensory rays and spicules. On the other hand, iff-1 mRNA is expressed in the gonad, and the lack of iff-1 activity causes sterility with an underproliferated germline resulting from impaired mitotic proliferation in both hermaphrodites and males. In spite of underproliferation, meiotic nuclei are observed, as revealed by presence of immunoreactivity to the anti-HIM-3 antibody; however, no gametogenesis occurs in the iff-1 gonads. These phenotypes are in part similar to the mutants affected in the components of P granules, which are the C. elegans counterparts of germ granules [Curr. Top Dev. Biol. 50 (2000) 155]. We found that localization of the P-granule component PGL-1 to P granules is disrupted in the iff-1 mutant. In summary, the two C. elegans homologues of eIF-5A act in different tissues: IFF-2 is required in the soma, and IFF-1 is required in the germline for germ cell proliferation, for gametogenesis after entry into meiosis, and for proper PGL-1 localization on P granules.
The developmental timing of all types of cells must be synchronized and spatially coordinated to achieve the organized development of a multicellular organism. Previously, we found RNAi of asb-1, encoding a germline-specific isoform of mitochondrial ATP synthase b subunit, caused 100% penetrant sterility in Caenorhabditis elegans. ATP synthase is one of the five complexes of the mitochondrial respiratory chain, and defects in some of the components of the chain are known to slow the growth and extend the lifespan of worms. We found that development of asb-1 mutant germ line was not arrested at any stage, but did slow to half the rate of wild type, whereas the rate of somatic development was the same in asb-1 mutants as that of wild type, indicating that asb-1 is required to maintain the rate of germline development but has no effect on somatic development. Among ATP synthase subunit genes, RNAi of asg-1, encoding a germline-specific isoform of the g subunit, also caused asb-1-like sterility, indicating that some other germline-specific components are also required to maintain the rate of germline development. Both asb-1 and asg-1 are located on autosomes while they possess counterparts, asb-2 and asg-2, respectively, on X chromosome, which are both required for somatic development. Chromosomal locations of the genes may be the basis of the segregation of germline/somatic functions of each gene, as were demonstrated for other autosomal/X-linked duplicated gene pairs.
scite is a Brooklyn-based organization that helps researchers better discover and understand research articles through Smart Citations–citations that display the context of the citation and describe whether the article provides supporting or contrasting evidence. scite is used by students and researchers from around the world and is funded in part by the National Science Foundation and the National Institute on Drug Abuse of the National Institutes of Health.