Cells decide to enter a new division cycle in response to extracellular signals during the G1 phase of the cell cycle. The mechanisms that act upon the basic cell-cycle machinery to accomplish temporal control of cell division are poorly understood. We are using the highly defined formation of the C. elegans vulva as a model to study the regulation of cell-cycle entry during development. In wild-type worms, six vulval precursor cells (VPCs) are formed during the L1 stage and have the potential to adopt vulval cell fates. The VPCs remain quiescent until they undergo further divisions and differentiate during the late L3 stage. To identify genes necessary for arrest of the VPCs in L1 to L3 larvae, we have screened for mutations that allow additional cell divisions during this period of development. In gain-of-function mutants of
lin-12 Notch, all six VPCs adopt vulval cell fates which results in the formation of up to 6 pseudovulvae. In this genetic background, mutations that cause extra cell divisions of the VPCs result in additional pseudovulvae (Hong et al ., 1998). We have screened EMS-mutagenized
lin-12(gf) animals and have identified two loci whose mutation yields the enhancer of
lin-12(gf) multivulva phenotype.   Homozygous
he118 mutants have extra cell divisions within the VPC lineage during the L2 stage, resulting in up to twice the wild-type number. The
he118 mutation has been mapped close to the
cki-1 gene, which encodes a cyclin-dependent kinase inhibitor. The phenotype of the
he118 mutant is rescued by a cosmid containing
cki-1 . In addition, RNA-mediated interference (RNAi) of
cki-1 also results in extra VPC divisions. Moreover, the extra VPC phenotype of both
he118 and
cki-1(RNAi) is enhanced by loss of
lin-35 Rb activity (See abstract by Boxem et al .). Based on these overlapping phenotypes and the map position of
he118 , we believe
he118 is a
cki-1 mutation. However, the
cki-1 coding region was found to be wild type in
he118 mutants. Since this mutation mainly affects the VPCs and, in contrast to
cki-1(RNAi) animals, not the intestinal, gonadal or other hypodermal lineages, the
he118 mutation may disrupt a tissue specific element required for proper
cki-1 expression.   In addition to
he118 we obtained two alleles,
he117 and
he119 , of
lin-1 Ets in the screen for extra cell divisions.
lin-1(lf) results in low penetrance extra VPC divisions during the L2 stage. Ectopic VPC divisions have also been observed in mutants of
lin-31 HNF-3, the presumed partner of LIN-1 in the VPCs (Miller et al., 1993). Using a
cki-1::GFP transgene (a kind gift from V. Ambros) we determined that
cki-1 expression is reduced or absent from the VPCs of
lin-1 or
lin-31 mutant animals. These data suggest that the extra VPC divisions in the
lin-1 and
lin-31 mutants may be a result of decreased expression of the cell cycle inhibitor
cki-1 . We propose that cell-type specific transcriptional regulation of the
cki-1 CDK inhibitor mediates the temporal withdrawal from the cell cycle during development.