Independent roles of a nuclear receptor gene in synapse specification and male mating in Caenorhabditis elegans

Finite numbers of genes generate organisms that are infinitely complex. Two solutions to this paradox are alternative splice products and pleiotropic expression patterns (single genes contributing to multiple characteristics). Here we show that in the nematode Caenorhabditis elegans the orphan nuclear receptor gene unc-55 exhibits both alternative splicing and pleiotropic expression as it contributes to two motor behaviors: locomotion and male mating. In C. elegans, the dorsal D (DD) and ventral D (VD) motorneurons (mns), referred to collectively as the D mns, compose a cross-inhibitory network. They share a number of structural and functional features, but are distinguished from one another by their synaptic patterns. The similarities are due to the expression of UNC-30, a transcription factor that activates structural and functional genes in both classes. A second transcription factor, UNC-55, which is a nuclear receptor belongs to COUP-TF subfamily, expressed in the VD mns, is necessary for generating the VD mn specific synaptic pattern. In unc-55 mutants the VD mns adopt the DD mn synaptic pattern. Conversely, ectopic expression of unc-55 in the DD mns results in their adoption of the VD mn synaptic pattern. Thus it appears that UNC-55 acts as a developmental switch that establishes the class specific synaptic patterns. An analysis and manipulation of the promoter region of a gene targeted by both UNC-30 and UNC-55 provide insight into the transcriptional events that create the similarities between the two functionally related neural classes while generating distinctive synaptic patterns. Beside unc-55 functions in VD mns, it is also necessary for male mating in C. elegans as indicated by the inability of male mating in unc-55 mutants. Most of the uncoordinated mutants in C. elegans that are unable to mate have severe locomotion defects or in addition have abnormal genital development. Is the inability to mate in unc-55 mutants due to the locomotion defect; or is it due to defects independent of locomotion? unc-55 has a sexually dimorphic expression pattern. Transgenic males express Punc-55: :gfp in a male specific neuron and structure cells associated with the spicules, a pair of appendages inserted into the vulva of the hermaphrodite during mating. Developmentally staged RNA interference experiments showed that adult expression of UNC-55 is required for successful copulation but expression in the motor neurons is not. There are two mRNA isoforms ( unc-55a and unc-55b) detected throughout postembryonic development in males, whereas only unc-55a is detected in hermaphrodites. The difference between the two isoforms is the presence of 13 amino acids that link the DNA binding domain to the ligand-binding domain. Two fusion genes were constructed one from each of the isoforms, both under the control of the unc-55 promoter and both with the gfp cassette on the 3' end. Although the gfp expression pattern in the motor neurons was the same for both, locomotion defects were rescued only by UNC-55A, the isoform expressed in both males and hermaphrodites. Although there were subtle differences in the gfp expression pattern; both isoforms were capable of rescuing male mating in an unc-55 mutant background. unc-55b appears to sustain male mating ability as they age. An unc-55 genetic pathway was also characterized to show that unc-55 expression in male tail but not VNC is regulated by a homeobox gene.