The Wnt/MAPK Signaling Pathway Controls Temporal Identities Of Seam Cells In C. Elegans

A cascade of heterochronic genes have been identified that form a complex regulatory pathway in controlling the temporal sequence of seam cell developmental events. Mutations in heterochronic genes cause stage-specific seam cell fates to be omitted or reiterated. The mechanisms by which signal transduction pathways are integrated with temporal information to ensure the adoption of distinct cell fate at a specific developmental stage in multicellular organisms remain largely unexplored.In a genetic screen to identify mutants which can suppress the retarded phenotype of dcr-1(bp132), we isolated a new allele of lit-1(bp239). In this study, we demonstrate that lit-1, encoding the Nemo-like kinase in Wnt/MAPK signaling, plays a role in specifying temporal identities of seam cells. The Wnt signaling pathway regulates multiple aspects of the development of stem cell-like epithelial seam cells in C. elegans, including cell fate specification and symmetric/asymmetric division. Loss of function of lit-1 suppresses defects in a class of retarded heterochronic mutants that show a reiteration of L2 stage events, while enhancing defects in precocious heterochronic mutants. Overexpressing lit-1 causes heterochronic defects opposite to those in lit-1 (lf) mutants.LIT-1::GFP localized to both cytoplasm and nucleus and was broadly expressed in a variety of cell types at post-embryonic stages, including in seam cells, vulva cells, neurons and pharyngeal muscles. lit-1 exhibits a periodic expression pattern in seam cells within each larval stage. The bp239 mutation result in the LIT-1 can not express in the seam cells at any stage,which is consistent with lit-1 (bp239) causes defects in the development of seam cells but does not result in other defects such as embryonic lethal associated with lit-1(null). The ceh-16::lit-1cDNA::gfp which expressed using a seam cell-specific ceh-16 promoter, displayed the same periodic expression pattern in seam cells as the above described lit-1::gfp reporter, indicating that the dynamic expression of LIT-1 is likely to be regulated at the post-transcriptional level.ceh-16::lit-1cDNA::gfp which only express the LIT-1 protein in seam cells slao can rescues the seam cell phenotype in dcr-1 lit-1 mutants, indicating that lit-1 functions cell-autonomously to regulate temporal fates of seam cells.The kinase activity of LIT-1 is required for its role in the heterochronic pathway. LIT-1b(K97G) mutantion in the predicted ATP binding site reslut in the LIT-1 protein loss its kinase activity. The ceh-16::lit-1b(K97G)::gfp reporter was expressed in seam cells at a level similar to that of ceh-16::lit-1::gfp, but it is not functional in rescuing the seam cell phenotype in dcr-1 lit-1 mutants. Thus, the kinase activity of LIT-1 is required for its role in suppressing the retarded heterochronic phenotype of dcr-1 mutants.Genetic analysis show that lit-1 (bp239) acts at a step downstream of miRNA biogenesis in dcr-1 mutants. Further study indicate that lit-1 specifies the temporal fate of seam cells is likely to through modulating miRNA-mediated silencing of target heterochronic genes. We further show that loss of function of other components of Wnt/MAPK signaling, including mom-4, wrm-1, apr-1 and pop-1, also causes heterochronic defects in sensitized genetic backgrounds.In a word, our study reveals a novel function of Wnt signaling in controlling the timing of seam cell development in C. elegans.In this genetic screen, we also identify a new allele of lin-4(bp238) which can dominantly suppressed the retarded heterochronic phenotype of dcr-1 mutants, and an allele of lin-42(bp245). The molecular understanding of autophagy has come almost exclusively from yeast genetic studies and referenced to mammalian system. Little is known about essential autophagy components specific to higher eukaryotes. Here we perform genetic screens in C. elegans to identify mutants with a defect in degradation of SEPA-1::GFP. From?10,000 genomes screened, we also isolated 9 mutant alleles of one novel genes, called epg-5. In epg-5 mutants, other autophagy targets, sunch as GFP::PGL-1, PGL-3 and T12G3.1::GFP also formed aggregates. Endogenous PGL granule components and T12G3.1, revealed by immunostaining assay, also formed aggregates in epg-5 mutant.In C. elegans, epg-5 encodes a novel autophagic protein C56C10.12. EPG-5 is highly conserved in higher eukaryotes, but is absent in yeast. epg-5 homolog of human KIAA1632 was identified in human breast cancers as a candidate cancer gene that is frequently mutated in breast cancers.epg-5::gfp which is functional in rescuing the defect in degradation of PGL granules in epg-5 mutants, is widely expressed. During embryogenesis. EPG-5::GFP was diffusely localized in the cytoplasm. At post-embryonic stages, epg-5::gfp was expressed in pharyngeal, body wall muscles and intestine cells.overexpression of epg-5 accelerates autophagic degradation in mutants with partial loss of function of genes involved in autophagosome expansion, indicating that epg-5 are rate-limiting factors in the autophagy pathway.