Molecular cloning and characterization of the dpy-20 gene of Caenorhabditis elegans

Mol Gen Genet. 1995 May 10;247(3):367-78. doi: 10.1007/BF00293205.

Abstract

We describe the molecular analysis of the dpy-20 gene in Caenorhabditis elegans. Isolation of genomic sequences was facilitated by the availability of a mutation that resulted from insertion of a Tc1 transposable element into the dpy-20 gene. The Tc1 insertion site in the m474::Tc1 allele was identified and was found to lie within the coding region of dpy-20. Three revertants (two wild-type and one partial revertant) resulted from the excision of this Tc1 element. Genomic dpy-20 clones' were isolated from a library of wild-type DNA and were found to lie just to the left of the unc-22 locus on the physical map, compatible with the position of dpy-20 on the genetic map. Cosmid DNA containing the dpy-20 gene was successfully used to rescue the mutant phenotype of animals homozygous for another dpy-20 allele, e1282ts. Sequence analysis of the putative dpy-20 homologue in Caenorhabditis briggsae was performed to confirm identification of the coding regions of the C. elegans gene and to identify conserved regulatory regions. Sequence analysis of dpy-20 revealed that it was not similar to other genes encoding known cuticle components such as collagen or cuticulin. The dpy-20 gene product, therefore, identifies a previously unknown type of protein that may be directly or indirectly involved in cuticle function. Northern blot analysis showed that dpy-20 is expressed predominantly in the second larval stage and that the mRNA is not at all abundant. Data from temperature shift studies using the temperature-sensitive allele e1282ts showed that the sensitive period also occurs at approximately the second larval stage. Therefore, expression of dpy-20 mRNA and function of the DPY-20 protein are closely linked temporally.

Publication types

  • Comparative Study
  • Research Support, Non-U.S. Gov't

MeSH terms

  • Amino Acid Sequence
  • Animals
  • Base Sequence
  • Caenorhabditis elegans / genetics*
  • Caenorhabditis elegans / metabolism
  • Caenorhabditis elegans Proteins*
  • Chromosome Mapping
  • Cloning, Molecular
  • Genes, Helminth*
  • Genomic Library
  • Genotype
  • Helminth Proteins / biosynthesis
  • Helminth Proteins / genetics*
  • Kinetics
  • Molecular Sequence Data
  • Mutagenesis
  • Phenotype
  • Recombinant Proteins / biosynthesis
  • Sequence Homology, Amino Acid
  • Sequence Homology, Nucleic Acid
  • Species Specificity
  • Temperature
  • Time Factors

Substances

  • Caenorhabditis elegans Proteins
  • Helminth Proteins
  • Recombinant Proteins
  • dpy-20 protein, C elegans

Associated data

  • GENBANK/U01918