ABSTRACT
Epithelial cells are polarized, with apical and basal compartments demarcated by tight and adherens junctions. Proper establishment of these subapical junctions is critical for normal development and histogenesis. We report the characterization of the gene let-413 which has a critical role in assembling adherens junctions in Caenorhabditis elegans. In let-413 mutants, adherens junctions are abnormal and mislocalized to more basolateral positions, epithelial cell polarity is affected and the actin cytoskeleton is disorganized. The LET-413 protein contains one PDZ domain and 16 leucine-rich repeats with high homology to proteins known to interact with small GTPases. Strikingly, LET-413 localizes to the basolateral membrane. We suggest that LET-413 acts as an adaptor protein involved in polarizing protein trafficking in epithelial cells.
Subject(s)
Caenorhabditis elegans Proteins , Caenorhabditis elegans/cytology , Caenorhabditis elegans/embryology , Cell Polarity , Epithelial Cells/cytology , Helminth Proteins/metabolism , Intercellular Junctions/metabolism , Amino Acid Motifs , Amino Acid Sequence , Animals , Caenorhabditis elegans/genetics , Caenorhabditis elegans/metabolism , Cell Adhesion , Cell Membrane/chemistry , Cell Membrane/metabolism , Cytoskeleton/metabolism , Cytoskeleton/ultrastructure , Epithelial Cells/metabolism , Epithelial Cells/ultrastructure , Epithelium/abnormalities , Epithelium/metabolism , Epithelium/ultrastructure , Genes, Helminth/genetics , Helminth Proteins/chemistry , Helminth Proteins/genetics , Intercellular Junctions/chemistry , Intercellular Junctions/ultrastructure , Microscopy, Electron , Molecular Sequence Data , Mutation/genetics , Phenotype , Protein Structure, Tertiary , Sequence AlignmentABSTRACT
In nematodes, flies, trypanosomes, and planarians, introduction of double-stranded RNA results in sequence-specific inactivation of gene function, a process termed RNA interference (RNAi). We demonstrate that RNAi against the Caenorhabditis elegans gene lir-1, which is part of the lir-1/lin-26 operon, induced phenotypes very different from a newly isolated lir-1 null mutation. Specifically, lir-1(RNAi) induced embryonic lethality reminiscent of moderately strong lin-26 alleles, whereas the lir-1 null mutant was viable. We show that the lir-1(RNAi) phenotypes resulted from a severe loss of lin-26 gene expression. In addition, we found that RNAi directed against lir-1 or lin-26 introns induced similar phenotypes, so we conclude that lir-1(RNAi) targets the lir-1/lin-26 pre-mRNA. This provides direct evidence that RNA interference can prevent gene expression by targeting nuclear transcripts. Our results highlight that caution may be necessary when interpreting RNA interference without the benefit of mutant alleles.
Subject(s)
Caenorhabditis elegans Proteins , Caenorhabditis elegans/genetics , Nuclear Proteins , Operon , RNA Precursors/genetics , RNA, Helminth/genetics , RNA, Messenger/genetics , Animals , Caenorhabditis elegans/growth & development , Carrier Proteins/genetics , DNA-Binding Proteins/genetics , Gene Deletion , Genes, Helminth , Helminth Proteins/genetics , Introns , Phenotype , Transcription Factors/genetics , Zinc FingersABSTRACT
The Caenorhabditis elegans LIN-26 protein is required to specify and/or maintain the fates of all non-neuronal ectodermal cells. Here we show that lin-26 is expressed until the somatic gonad primordium stage in all cells of the somatic gonad, except in distal tip cells, and later in all uterine cells. To determine if lin-26 functions in the somatic gonad, we have generated gonad-specific lin-26 alleles obtained by integration of lin-26 promoter deletion derivatives into a lin-26 null mutant background. In this way, we rescued the lethal phenotype imparted by lin-26 null mutations and uncovered a highly penetrant sterile phenotype. Specifically, the strongest of these new alleles was characterized by the absence of lin-26 expression in the somatic gonad, the presence of endomitotic oocytes, decreased germline proliferation, a protruding vulva and a less penetrant absence of gonad arms. Lineage analysis of mutant somatic gonads and examination of several markers expressed in the spermatheca, sheath cells, distal tip cells and the uterus, suggest that LIN-26 is required in sheath, spermatheca and uterine precursors, and in uterine cells. We conclude that lin-26 performs a similar function in the non-neuronal ectoderm and the somatic gonad, a mesoderm derivative, and we speculate that lin-26 is required to express epithelial characteristics.
Subject(s)
Caenorhabditis elegans Proteins , Caenorhabditis elegans/genetics , DNA-Binding Proteins/physiology , Gonads/growth & development , Transcription Factors/physiology , Animals , Animals, Genetically Modified , Female , Fluorescent Antibody Technique , Gene Expression Regulation, Developmental/genetics , Genes, Helminth/genetics , Male , Oocytes/pathology , Pedigree , Promoter Regions, Genetic/genetics , Sequence Deletion/geneticsABSTRACT
The mutation lin-26(n156) prevents vulva formation in C. elegans by transforming the vulval precursor cells into neurons or neuroblasts. We have isolated and characterized three new lin-26 alleles, which result in embryonic lethality. These mutations cause a few other hypodermal cells to express a neural fate and most hypodermal cells to degenerate. lin-26 encodes a presumptive zinc-finger transcription factor. Our data indicate that lin-26 is required for cells to acquire the hypodermal fate.
