Worm Breeder's Gazette 10(3): 109

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.

Genes and Proteins of the Muscle Dense-Bodies

Robert Barstead, Lawrence Kleiman and Robert Waterston

In the body wall muscle of the nematode actin filaments are anchored 
to the sarcolemma through the dense-bodies.  Two of the protein 
components of the dense-bodies have been characterized using 
monoclonal antibodies (Francis and Waterston, 1985).  One of these 
components, p107b/110, is found at the base of the dense-bodies 
adjacent to the sarcolemma and, as we have previously reported, the 
deb-1 gene which encodes this antigen has been cloned from an 
expression library.  We have completed the sequencing of this gene and 
an associated cDNA, and have now shown that it encodes a peptide that 
is highly similar to chicken vinculin.  We have further shown that the 
size difference between chicken vinculin, Mr 130,000, and its nematode 
analog, Mr 107,000, is due in part to the absence from the nematode 
sequence of one of the three internal 110 amino acid repeats found in 
the chicken sequence.  While the function of vinculin is not known, 
its location in structures which mediate the attachment of actin 
filaments to membranes as well as its biochemical properties has led 
to the speculation that it is important for the initiation of actin 
filament assembly.  We are currently attempting to recover mutations 
in the deb-1 gene which should allow us to assess the developmental 
role of nematode vinculin.
A second component of the dense-body, p107a, has been shown to be 
immunologically similar to the actin binding protein alpha-actinin.  
After failing to recover the gene for p107a using antibodies to screen 
an expression library, we decided to make use of two known alpha-
actinin cDNA sequences from dictyostelium and chicken.  Degenerate 
oligonucleotide probes to the conserved actin binding region of the 
known sequences were synthesized and used to screen a genomic library. 
A set of overlapping clones was recovered and sent to Cambridge where 
the clones were mapped to a single locus on chromosome V on the same 
contig as myo-3, the gene encoding the minor heavy chain myosin of the 
body wall muscle.  Other than myo-3, there are no known muscle 
affecting genes in this region.  We are currently sequencing a 3.8 kb 
cDNA for the gene.  The sequence obtained thus far shows extensive 
similarity with other known alpha-actinin sequences.  Moreover, when 
portions of the cDNA clone are fused to the E.  coli  -gal gene, a 
fusion protein is made which reacts with antibodies that recognize 
p107a.  We are currently focusing on completing the sequence of the 
cDNA.  Now that both the alpha-actinin and the vinculin genes are 
cloned, we are in a position to genetically dissect the interactions 
between these two proteins and their role in the initiation of thin 
filament assembly.