The
let-23 locus was originally defined by three larval lethal alleles [Herman, 1980; Sigurdson et al., 1984] and one cold-sensitive semi-lethal vulvaless allele,
n1045 [Ferguson & Horvitz, 1985]. Our interest in
let-23 was sparked by the observation that
n1045 causes a novel phenotype at 25 C -- hyperinduction of the vulva, in which four or even five of the vulval precursor cells (VPCs) are induced. (In wild type, three of six VPCs are induced; in Vulvaless (Vul) animals, zero of six are induced.) This Hyperinduced or Hin phenotype requires a signal from the anchor cell and thus is distinct from the classic Multivulva (Muv) phenotype. That
let-23 could be mutated to give opposite phenotypes --
n1045 at 15 C is Vul, but at 25 C is Hin -- led us to believe that
let-23 plays a critical role in the specification of VPC fate in response the inductive signal from the anchor cell. We are therefore genetically and molecularly analyzing the
let-23 locus. Our first genetic screen to obtain new
let-23 alleles made use of the fact that
let-23 is the only 'Determination' gene known to suppress the Muv phenotype of
lin-15(
n309) [Ferguson et al., 1987]. We therefore EMS-mutagenized
lin-15(
n309) and looked in the F2 for phenotypic revertants that result in a Vulvaless (Vul) phenotype when crossed away from
n309. Using this screen, we isolated
sy1, which maps to the
let-23 locus and fails to complement
n1045 for the Vul phenotype. 93% of homozygous
sy1 hermaphrodites are egglaying- defective, a result of the Vul phenotype. All
sy1 homozygotes are viable, and
sy1 complements all known
let-23 alleles for the Lethal phenotype. From additional
lin-15 reversion screens, we have obtained another Vul mutation linked to
let-23.
n1045 is recessive and results in a Vul phenotype in trans to a deficiency for the locus at all temperatures, suggesting that
n1045 results in a decreased level of
let-23 activity at all temperatures. There are two reasonable hypotheses for the Hin phenotype.
n1045 might be a neomorph that is dose-dependent. Alternatively, although levels are down with respect to wild type,
n1045 has more activity at 25 C than at 15 C, and a particular level of
let-23 activity might result in a Hin phenotype due to a bizarre dose-response curve. Complementation tests with
sy1 have provided evidence favoring the hypothesis that while a severe decrease in
let-23 activity results in a Vul phenotype, a less severe decrease in activity results in a Hin phenotype. The key observation is that the Hin phenotype can be recreated without using
n1045, using instead
sy1 and a lethal allele,
mn224. In particular, 20% of
sy1/mn224 heterozygotes are Hin; the remaining 80% are wild-type. We interpret the Hin phenotype as resulting from a failure to induce the 'Lateral Inhibitory Signal' [LIS] described in the last newsletter [Sternberg WBG 10(1)]. Specifically, we propose that in wild type the LIS is activated in VPCs in response to
let-23, which in turn is activated by the signal from the anchor cell, but at a higher level of
let-23 activity than required for generating vulval sublineages. In a Hin animal, we argue, VPCs are induced to generate vulval sublineages but fail to inhibit their neighbors from also participating in vulval development; hence more than three VPCs can generate vulval sublineages. Since
sy1/Df hermaphrodites are highly penetrant for the Vul phenotype but are nonetheless viable, we screened for new EMS-induced mutations that fail to complement
sy1. We have isolated fifteen new
let-23 alleles after screening 20,000 F1 chromosomes. Each mutation is tightly linked to
let-23 and fails to complement
sy1 and
n1045 for the Vul phenotype. Two of these alleles,
sy10 and
sy12, are incompletely penetrant lethals, causing 80% and 90% of larvae to die, respectively. The surviving worms show two new phenotypes: all hermaphrodites are sterile, and all males have short, crumpled spicules. The cellular bases of these phenotypes are under investigation because we are dying to know if
let-23 is involved in the specification of other cell fates besides the VPCs and P11/P12. We are pursuing two approaches to clone the
let-23 locus: we are screening for transposon-induced alleles that fail to complement
sy1, and will use the over 20 existing alleles to identify RFLPs within a chromosomal walk