Post-embryonic Expression of HLH-3 is Sexually Dimorphic and Affects Sexually Dimorphic Behavior in Males

A critical stage of neuronal development is the expression of proneural bHLH genes in the ectoderm leading to the specification of neuronal precursor cells. Expression of subsequent identity specification genes allow these cells to mature and acquire terminal identity features including a particular morphology, connectivity, and transcriptome. Other bHLH transcription factors are known to have alternate, non-traditional roles in differentiation. This work investigates the role that one such non-traditional bHLH transcription factor, hlh-3, plays in neuronal development. The goal of this work is to characterize the post-embryonic expression pattern of hlh-3 in males, as well as the differentiation of a subset of neurons required for male specific behaviors. Our findings show that expression of hlh-3 is sexually dimorphic in the L1, late L3/early L4, and adult stages. This work also describes the role of hlh-3 in the maturation of a subset of neurons required for male specific exploration behavior. Our findings show that in the absence of hlh-3, AWA, ASJ, and AIM neurons fail to acquire terminally differentiated features and therefore contribute to the failure of males to perform behaviors mediated by these cells. We further characterize features of another sex-shared neuron with sex-specific function, PHCs, including anterior axon length, eat-4/VGLUT relative expression, overall axon morphology, and male mating behavior to determine if hlh-3(lof) males have fully differentiated PHC neurons. Our results show that while eat-4/VGLUT levels and anterior axon length are unaltered, overall axon morphology is severely compromised, and subsequent male mating behaviors fail at multiple points. These results indicate that hlh-3(lof) male PHCs are likely to be a contributing factor to the large-scale defects observed in male mating behaviors