Multiple roles for the E/Daughterless ortholog HLH-2 during C. elegans gonadogenesis

AbstractHLH-2 is the Caenorhabditis elegans ortholog of the Drosophila Daughterless and mammalian E basic helix–loop–helix (bHLH) transcriptional activators that function during diverse events during animal development. HLH-2 has been implicated in cell fate specification in different neural lineages and in the LIN-12/Notch-mediated anchor cell (AC)/ventral uterine precursor cell (VU) decision in the somatic gonad. Here, we show that hlh-2 plays several distinct roles during somatic gonadogenesis. Our analysis suggests that hlh-2 is required to endow specific somatic gonadal cells with the competence to undergo the AC/VU decision, as well as functioning in the AC/VU decision per se; this novel “proAC” role appears to be analogous to the proneural role of Drosophila Daughterless. In addition to its two distinct roles in the specification of the AC, hlh-2 is also required for correct differentiation and function of the AC. hlh-2 also acts at an independent point in the gonadal lineage both to specify distal tip cells (DTCs) and in DTC differentiation and function

sel-11 and cdc-42, Two Negative Modulators of LIN-12/Notch Activity in C. elegans

Background: LIN-12/Notch signaling is important for cell-cell interactions during development, and mutations resulting in constitutive LIN-12/Notch signaling can cause cancer. Loss of negative regulators of lin-12/Notch activity has the potential for influencing cell fate decisions during development and the genesis or aggressiveness of cancer. Methodology/Principal Findings: We describe two negative modulators of lin-12 activity in C. elegans. One gene, sel-11, was initially defined as a suppressor of a lin-12 hypomorphic allele; the other gene, cdc-42, is a well-studied Rho GTPase. Here, we show that SEL-11 corresponds to yeast Hrd1p and mammalian Synoviolin. We also show that cdc-42 has the genetic properties consistent with negative regulation of lin-12 activity during vulval precursor cell fate specification. Conclusions/Significance: Our results underscore the multiplicity of negative regulatory mechanisms that impact on lin-12/ Notch activity and suggest novel mechanisms by which constitutive lin-12/Notch activity might be exacerbated in cancer

OrthoList: A Compendium of C. elegans Genes with Human Orthologs

C. elegans is an important model for genetic studies relevant to human biology and disease. We sought to assess the orthology between C. elegans and human genes to understand better the relationship between their genomes and to generate a compelling list of candidates to streamline RNAi-based screens in this model.We performed a meta-analysis of results from four orthology prediction programs and generated a compendium, "OrthoList", containing 7,663 C. elegans protein-coding genes. Various assessments indicate that OrthoList has extensive coverage with low false-positive and false-negative rates. Part of this evaluation examined the conservation of components of the receptor tyrosine kinase, Notch, Wnt, TGF-ß and insulin signaling pathways, and led us to update compendia of conserved C. elegans kinases, nuclear hormone receptors, F-box proteins, and transcription factors. Comparison with two published genome-wide RNAi screens indicated that virtually all of the conserved hits would have been obtained had just the OrthoList set (∼38% of the genome) been targeted. We compiled Ortholist by InterPro domains and Gene Ontology annotation, making it easy to identify C. elegans orthologs of human disease genes for potential functional analysis.We anticipate that OrthoList will be of considerable utility to C. elegans researchers for streamlining RNAi screens, by focusing on genes with apparent human orthologs, thus reducing screening effort by ∼60%. Moreover, we find that OrthoList provides a useful basis for annotating orthology and reveals more C. elegans orthologs of human genes in various functional groups, such as transcription factors, than previously described

Suppressors of the egg-laying defective phenotype of sel-12 presenilin mutants implicate the CoREST corepressor complex in LIN-12/Notch signaling in C. elegans

Presenilin is an essential component of the LIN-12/Notch signaling pathway and also plays a critical role in the genesis of Alzheimer's disease. Previously, a screen for suppressors of the egg-laying defective phenotype caused by partial loss of presenilin activity in Caenorhabditis elegans identified a number of new spr genes that are potentially involved in the regulation of LIN-12/Notch signaling or presenilin activity. Here we report the molecular identity of two spr genes, spr-1 and spr-5. Our genetic analysis indicates that loss of spr-1 elevates lin-12/Notch gene activity in many different cell fate decisions, suggesting that spr-1 is a negative regulator of LIN-12/Notch signaling. Sequence analysis revealed that spr-1 is an ortholog of human CoREST, a known corepressor. SPR-1 is localized to the nucleus and acts in a cell-autonomous manner; furthermore, human CoREST can substitute for SPR-1 in C. elegans. We also show that spr-5 encodes a homolog of p110b, another known member of the CoREST corepressor complex. Our results suggest that the CoREST corepressor complex might be functionally conserved in worms, and we discuss the potential role of SPR-1 and SPR-5 in the repression of transcription of genes involved in, or downstream of, LIN-12/Notch signal transduction

Determinants in the LIN-12/Notch Intracellular Domain That Govern Its Activity and Stability During Caenorhabditis elegans Vulval Development

Upon ligand binding, the LIN-12/Notch intracellular domain is released from its transmembrane tether to function in a nuclear complex that activates transcription of target genes. During Caenorhabditis elegans vulval development, LIN-12/Notch is activated by ligand in two of six multipotential vulval precursor cells (VPCs), specifying the “secondary vulval fate” and descendants that contribute to the vulva. If LIN-12 is ectopically activated in other VPCs, they also adopt the secondary fate, dividing to produce extra vulval cells, resulting in a “Multivulva” phenotype. Here, we identify determinants in the LIN-12 intracellular domain [“LIN-12(intra)”] that govern its activity and stability during C. elegans vulval development; we assayed activity of mutant forms based on their ability to cause a Multivulva phenotype and stability using a GFP tag to visualize their accumulation. Our analysis has revealed that, while the ubiquitin ligase SEL-10/Fbw7 promotes LIN-12(intra) downregulation in VPCs, there is a distinct mechanism for downregulation of LIN-12(intra) in VPC descendants. Our analysis also revealed that LIN-12(intra) must be in the nuclear complex to be regulated appropriately in VPCs and their descendants, and that the structure or conformation of the carboxy-terminal region influences stability as well. Although activity and stability are generally well-correlated, exceptions where they are uncoupled suggest that there may be roles for the carboxy-terminal region and sel-10 that are independent of their roles in regulating LIN-12(intra) stability

EMB-4: A Predicted ATPase That Facilitates lin-12 Activity in Caenorhabditis elegans

The sel-6 gene was previously identified in a screen for suppressors of the egg-laying defect associated with hypermorphic alleles of lin-12 (Tax et al. 1997). Here we show that sel-6 and two other previously defined genes, mal-2 and emb-4, are the same gene, now called “emb-4.” We perform a genetic and molecular characterization of emb-4 and show that it functions cell autonomously as a positive regulator of lin-12 activity. Viable alleles identified as suppressors of lin-12 are partial loss-of-function mutations, whereas the null phenotype encompasses a range of lethal terminal phenotypes that apparently are not related to loss of lin-12/Notch signaling. emb-4 encodes a large nuclearly localized protein containing a predicted ATPase domain and has apparent orthologs in fission yeast, plants, and animals