While both neurons and glia undergo sex-specific changes, most attention has focused on sex differences in neurons. In principle, glia could develop sex differences in response to their neuronal partners or they could have their own cell-autonomous programs for sexual differentiation. In C. elegans, we discovered that CEPso glia become sexually dimorphic and initiate expression of a transcriptional reporter (
grl-18pro:GFP) in adult males only. This change in CEPso glia occurs concomitantly with the activation of their neuronal partners, the male-specific CEM neurons. However, by genetically manipulating the sex identity of neurons and glia, we found that this sex-specific change in CEPso glia does not depend on CEM neurons but instead is determined by the sex identity of glia alone. Using candidate and unbiased genetic screens, we identified the timing factors
lep-2/Makorin and
lep-5 as well as the sex identity factor
mab-3/DMRT as required for the sexual differentiation of CEPso glia. In these mutants, adult male
grl-18 expression is delayed or absent, respectively. These factors have also been shown to control sexual differentiation of neurons, but we find that
mab-3 acts cell-autonomously in CEPso glia to switch on adult male-specific genes. Finally, we isolated three novel alleles of the transcriptional repressor
nfya-1/NF-Y, which has not previously been implicated in sexual differentiation of the nervous system. In
nfya-1 mutants, CEPso glia of adult hermaphrodites inappropriately express
grl-18. Epistasis analysis shows that loss of
nfya-1 completely bypasses the requirement for
mab-3, suggesting that
nfya-1 may be a novel downstream effector of
mab-3 that regulates sex-specific gene expression in glia. Together, our findings suggest a model in which male-specific expression of
mab-3 in CEPso glia leads to the repression of NFYA-1, thereby allowing male genes to switch on in adults. These results demonstrate that glia do not passively respond to changes in their neuronal surroundings but instead initiate their own program of sexual differentiation.