Worm Breeder's Gazette 10(2): 74

These abstracts should not be cited in bibliographies. Material contained herein should be treated as personal communication and should be cited as such only with the consent of the author.

Isolation and Characterization of Basement Membrane Collagen Genes in C. elegans

Xiaodu Guo and Jim Kramer

Basement membranes(BMs) are believed to be involved in tissue 
morphogenesis, maintenance of tissue architecture, as well as cell 
movement and attachment.  Although BM components have been studied 
intensively at the biochemical level, there has been no means to study 
the functions of these components genetically.  We are interested in 
identifying the genes that encode BM components and in performing 
molecular and genetic studies of the functions of BM in development.  
Using fragments of a mouse Type IV (BM) collagen cDNA as probes, we 
have identified two clones containing BM-like collagen genes (CH#1 and 
CH#2) from a C.  elegans genomic sublibrary of collagen containing 
phage.  The two genomic clones were isolated by screening the 
sublibrary with a fragment of the mouse Type IV collagen cDNA that 
does not contain any Gly-X-Y coding sequence and does not cross 
hybridize to other collagen genes.  Under relatively high stringency 
conditions, CH#1 and CH#2 hybridize more strongly to the Gly-X-Y 
portion of the mouse Type IV cDNA than do any of the other C.  elegans 
collagen clones.  CH#1 and CH#2 have been restriction mapped and both 
genes span about a 6-7 kb region within the clones.  The restriction 
maps are different from each other indicating that these two genes 
encode two distinct alpha polypeptide chains of Type IV collagen.  
Both clones were radioactive labeled and probed to Northern blots.  
The results show that both genes produce 5.5 kb long transcripts, 
similar in size to the mouse and human Type IV collagen transcripts, 
but much larger in size than the C.  elegans cuticle collagen 
CH#1 and CH#2 have been placed onto contigs by the MRC group.  The 
genetic location of the CH#1 contig is not known.  CH#2 has been 
mapped to linkage group III, about 100 kb to the right of the lin-12 
locus.  Several embryonic lethal genes are located in this region.  We 
have examined mutants of these genes briefly.  One of these genes, emb-
9, has temperature sensitive mutant phenotypes that could result from 
defects in BM.  Morphologically, emb-9 appears very similar to let-2, a 
gene previously found to cause BM defects (J.  Kramer and J.  Priess). 
Two of the five emb-9 alleles are semidominant, suggestive of a 
structural protein like a BM collagen.  emb-9 is being analyzed to 
determine if it is the CH#2 BM collagen gene, and this may provide the 
means to begin a genetic analysis of BM structure and function.