Here we describe the isolation and cloning of two EMS-induced mutations in the
cwn-2 gene in a forward genetic screen for mutants with defects in the position of the nerve ring. The screen was done in a
slt-1 mutant background to sensitize for genes functioning redundantly with
slt-1, and some of the
cwn-2 phenotypes are greatly enhanced by mutations in
slt-1.
slt-1 encodes a secreted ligand for the
sax-3(Robo) axon guidance receptor.
sax-3 mutants have defects in the position of the nerve ring in which the nerve ring is more anterior than wild type.
cwn-2 mutants also have anterior nerve rings, while
slt-1 mutant worms have no defect in nerve ring position. Double mutants in
cwn-2 and
sax-3 have defects more severe than either single mutant, indicating that
cwn-2 acts at least partly in parallel to
sax-3, and thus is not likely in the
sax-3 pathway.
cwn-2 encodes a member of the Wnt family of secreted glycoproteins, a family known to influence cell fate, migration and axon guidance. Both EMS alleles of
cwn-2 have CAN cell migration defects in addition to the nerve ring defects. They also have branching defects in the axon of the ADL neuron. A strain provided by the Gene Knockout Consortium which has a deletion in the
cwn-2 gene has similar defects. The CAN migration defect was independently noted by Zinovyeva and Forrester (2005). These defects are rescued by a transgene containing the
cwn-2 gene, establishing that these defects are caused by mutations in
cwn-2.
cwn-2 reporter genes are expressed in the pharynx, gut, rectal gland cells, and the anterior half of the body wall muscle, and in the SMD neurons. Expression of
cwn-2 in either the body wall muscle or the pharynx can rescue both the nerve ring and CAN migration defects. Results of over-expression and misexpression experiments will be presented.
A candidate gene approach was used to find potential receptors for
cwn-2. Worms mutant in
cam-1 have anterior nerve rings. Anterior nerve rings have also been reported in
cfz-2 mutants (Zinovyeva and Forrester, 2005).
cam-1 or
cfz-2 could possibly interact with
cwn-2 in nerve ring development. Double mutants in
cwn-2 and
cfz-2 are more severe than either single mutant, indicating that they have some parallel functions, and that another Wnt may be acting through
cfz-2 to affect nerve ring position.