During development of the hermaphrodite vulva, three out of six equivalent Pn.p cells in the ventral hypodermis are induced by a signal from the neighbouring gonadal anchor cell (AC) to adopt vulval cell fates. The AC signal is transduced in P6.p by an evolutionary conserved RTK/RAS/MAP kinase pathway where it specifies the primary cell fate. P6.p then induces its neighbours P5.p and P7.p through a lateral signal to adopt the secondary cell fate. While many genes required for activation of the RTK/RAS/MAPK pathway have been characterized, less is known about genes that negatively regulate this signaling pathway. It has been proposed that members of the CL100/PAC-1/MKP-3 family of dual-specificity phosphatases negatively regulate MAP kinases. The C. elegans genome encodes about 185 putative phosphatases. Among these genes, C05B10.1 encodes the closest homolog of CL100/PAC-1/MKP-3 and we refer to this gene as
mkp-1. To investigate the role of
mkp-1 during vulval development, we have isolated an EMS-induced deletion of bp (
mkp-1(
zh15)) that removes exons 2 to 6 which encode the putative MAP kinase (MPK-1) binding site.
mkp-1(
zh15) single mutants exhibit a wild-type vulval phenotype. However,
mkp-1 (
zh15) strongly suppresses the vulvaless (Vul) phenotype caused by mutations that reduce but do not eliminate the activity of the AC signaling pathway (eg.
let-23 (
sy1),
lin-2(
n397),
lin-7(
e1413) and
sem-5(
n2019)). Furthermore, over-expression of
mkp-1 in the Pn.p cells under control of the Pn.p cell-specific
lin-31 promoter (
lin31-
mkp-1) causes a dominant Vul phenotype. Taken together, these data indicate that
mkp-1 negatively regulates vulval induction. In addition to the genetic studies, we have examined the expression pattern of
mkp-1 by generating
mkp-1::GFP reporter transgenes. MKP-1::GFP is widely expressed throughout the embryo and in most somatic cells during larval development. In particular, in early L2 animals all Pn.p cells express MKP-1::GFP. However, in late L2/ early L3 animals, when the VPC fates are determined,
mkp-1::GFP expression is down-regulated in P6.p but persists in P5.p and P7.p. Finally, we observed that introduction of a nuclear localization signal into the
mkp-1::GFP reporter construct (
mkp-1::NLS::gfp) results in a penetrant Vul phenotype, similar to the Vul phenotype observed in
lin31-
mkp-1 animals. This observation suggest that
mkp-1 may act predominantly in the nucleus of the Pn.p cells. In summary, our results support the idea that wild-type
mkp-1 may function as an inhibitor of the RTK/RAS/MAPK signaling pathway by negatively regulating MPK-1 activity. Currently, we are performing further genetic studies and biochemical experiments to test this hypothesis.