UBC Theses and Dissertations
A genetic and molecular analysis of the bli-4 locus of Caenorhabditis elegans, an essential gene encoding KEX2-like subtilisin-typw serine endoproteases Peters, Kenneth W.
This thesis reports a characterization of genetic mutations and molecular structure of the bli-4 (I) gene of Caenorhabditis elegans. The bli-4locus had been previously defined by a single recessive mutation, e937, which disrupts the structure of adult-stage cuticle causing the formation of fluid-filled separations of the cuticle layers, or blisters. A characterization of e937 and eleven additional mutations is reported. The mutations were grouped into three classes based on phenotype and complementation analysis: Class I, represented by the single allele causing blistering, e937;Class II, nine lethal mutations that arrest development at the end of embryogenesis and that fail to complement all other mutations; and Class 111,two larval lethal mutations, s90 and h754, that complement e937. The complementation pattern provides evidence that all of these mutations are allelic, and that bli-4 is a complex locus with an essential function late in embryogenesis. A region of chromosome I that includes the bli-4 locus was identified by aligning the C. elegans genetic and physical maps. The bli-4 coding region was identified by using cosmids as hybridization probes to detect chromosomal alterations in the DNA of bli-4 mutant strains. Two mutations of bli-4 are small rearrangements of the gene; e937, is a 3.5 kilo base (kb)deletion, and h1010 is an insertion of the 1.6 kb transposable element, Tc1.Three protein products that differ at the carboxyl end, are predicted from alternately spliced bli-4 cDNA clones. The predicted proteins were designated blisterin A, blisterin B and blisterin C according to the order that the variantly spliced 3' ends occur on the chromosome. Of the three bli-4cDNA clones characterized, only blisterin B includes an open reading frame beginning with an ATG start codon. The blisterin A and blisterin C open reading frames begin within blisterin B, and are likely to be incomplete. The predicted blisterin B gene product has a potential secretion signal peptide atits amino terminal. Blisterin C has a potential Tran membrane domain near its carboxyl terminal while blisterin A and B lack this domain. The blisterins share significant sequence identity with kex2-like serine endoproteases ,which are responsible for the cleavage of secreted proteins in yeast and mammals. This characterization of bli-4 provides the first evidence for an essential role of a KEX2-like gene in the development of a multi-cellular organism.
Item Citations and Data