fem-3 specifies male fates in C. elegans. In hermaphrodites, gain-of-function (gf)
fem-3 mutants fail to switch from spermatogenesis to oogenesis. The molecular defects found in
fem-3(gf) mutations affect a 5 bp region within the
fem-3 3'UTR and several lines of evidence suggest that this regulation acts at the level of translation. Our working hypothesis is that the
fem-3 3'UTR possesses a negative cis-acting translational control element, which may include the 5 bp region altered in
fem-3 (gf) mutants. We have developed a reporter assay to examine this 3'UTR-mediated translational control of
fem-3. Two types of constructs have been made: lacZ::
fem-3(+) 3'UTR has lacZ coding sequences fused to the wildtype
fem-3 3'UTR, while lacZ::
fem-3(gf) 3'UTR has lacZ coding sequences fused to a
fem-3 (gf) 3'UTR. Both reporter transgenes are under the control of a heat shock promoter. Hermaphrodites containing the lacZ::
fem-3(+) 3'UTR transgene show undetectable levels of B-gal activity (assayed by x-gal staining) whereas worms that contain the lacZ::
fem-3(gf) 3'UTR show high levels of B-gal activity. Despite these dramatic differences in B-gal activity, both transgenic strains possess equal levels of reporter mRNA, as determined by RNAse protection. These results strongly support the idea that the
fem-3 3'UTR carries a cis-acting translational regulatory element. Furthermore, this
fem-3 3'UTR regulation is not limited to the germline: expression of the reporter transgene is observed in multiple somatic tissues. Somatic regulation of the
fem-3 3'UTR was predicted due to somatic affects observed in
tra-1(gf);
fem-3(gf) double mutants (Schedl and Kimble, 1988). We are now using this assay to identify genes that when mutant, activate the lacZ::
fem-3 (+) 3'UTR reporter transgene. Our results will be reported at the meeting.