A large gene family first defined in C. elegans by
unc-7 and
eat-5, and in Drosophila by Shaking-B/Passover and l(1)ogre, represents an invertebrate equivalent of the vertebrate gap junction proteins, or connexins. Although the two gene families share no significant sequence similarity, the proteins from these two families are functionally similar. Recently it has been shown that the Drosophila Shaking-B protein, an essential protein of the Shak-B/Pas locus, can mediate electrical coupling in paired Xenopus oocytes, the defining assay for connexin function (Phelan et al., Nature 391:181, 1998). Since at least one of these invertebrate proteins is capable of forming channels, it has been proposed that these proteins be referred to as innexins, for invertebrate connexins. Our previous collaborative efforts to show coupling in Xenopus oocytes with worm innexins failed. Attempts to show novel channel formation in insect cell lines expressing a worm innexin also failed. Recently Y. Landesman and D. Paul have been able to demonstrate electrical coupling in paired Xenopus oocytes mediated by one of our worm innexin constructs. With the function of this gene product now established, we refer to this innexin gene as
wxn-1 (worm innexin). We are particularly interested in the role of gap junctions in determining the specificity of nervous system wiring, and the role of gap junctions during development. We are studying
unc-7 to address questions concerning wiring in the ventral nerve cord.
unc-7(
e5) animals appear to possess ectopic gap junctions (J. White et al.), and the sequence of
e5 predicts an early stop codon, making
e5 a likely null allele. Therefore we postulate that UNC-7 is involved in the specificity of gap junction formation. We have generated specific antibodies to the carboxyl terminus of UNC-7, and these antibodies detect UNC-7 in the nerve ring and the ventral cord. Because of the punctate nature of the staining pattern, it is not possible to identify individual cells expressing UNC-7. To investigate the function of gap junctions during development, we are studying
wxn-1. We have generated antibodies to the carboxyl terminus of WXN-1, and these antibodies confirm and elaborate on the expression pattern seen using WXN-1::GFP fusion constructs. WXN-1 is seen at the two cell stage, and is distributed in small punctate aggregates found exclusively along regions of cell contact. The protein is expressed throughout early embryogenesis in apparently all cells. As morphogenesis proceeds, WXN-1 becomes more restricted in distribution, and by the time of hatching is most clearly present in the pharynx and hypodermal cells. The protein is expressed transiently in developing motorneurons in the ventral nerve cord at late L1. WXN-1 is also expressed in the developing sex muscles into adulthood. In an attempt to isolate mutations in
wxn-1, we carried out a screen for lethals rescued by expression of
wxn-1 on an extrachromosomal array. Two candidate mutations were isolated. One is a lethal which maps to the same region as
wxn-1. The majority of these mutants die just prior to or shortly after hatching. Occasional escapers arise, which are short, slow-growing, are sterile or have few progeny, and display some hypodermal defects. Sequencing of the
wxn-1 coding region failed to detect a mutation. Antibody studies suggest that WXN-1 protein is present , but levels may be reduced. Therefore the mutation may lie in a regulatory region of
wxn-1, or in a second gene affecting
wxn-1 expression that maps nearby. The second mutation isolated in the screen maps outside the
wxn-1 region, to a region with another worm innexin gene. We are currently trying to determine if this other innexin and
wxn-1 may have some partially redundant functions.