We have recently characterised differentiated glial cells that divide to produce neurons, the mystery cells of the male (MCMs). Here we present a previously undescribed direct glia-to-neuron cell fate switch, revising the total number of male neurons to 387 and glia to 90. Studies of vertebrate neural development have revealed that differentiated glia can act as neural precursors, however the cellular and molecular mechanisms have not been fully determined. To identify the genetic factors that regulate these glia-to-neuron cell fate switches, we have generated a collection of no mystery cell (nom) mutants and assessed their role in both glia-to-neuron cell fate switches. In early L4, the male amphid socket (AMso) glial cells divide asymmetrically. We observe increased nuclear localization of the Wnt-signalling effector POP-1/TCF, in the anterior daughter, the self-renewed AMso than in the posterior MCMs. The proneural factor
hlh-14/Ascl1 is then transiently expressed in the MCMs, prior to neuronal differentiation. From a GFP-based forward genetic screen of 4000 genomes we have isolated nine nom mutants in which the MCMs fail to be specified. We have identified mutants that affect AMso division, MCM neuronal differentiation and MCM neuronal subtype specification. Two AMso division mutants,
nom-5 and
nom-8 were mapped-by-sequence to the
cdk-4 locus. CDK-4 is required for G1-S progressions in postembryonic blast cells but surprisingly, our alleles only appear to affect the AMso division. Moreover, no loss of glial fate or acquisition of neuronal characteristics was observed in the undivided AMso, suggesting that MCM specification requires DNA replication or the asymmetric segregation of neural factors. Following Sulston's preliminary observations, we find that the male phasmid socket one (PHso1) glial cells become bona fide neurons during the L4 stage and name them phasmid neuron D (PHD). Importantly, this transition is direct, not requiring cell division. We establish their developmental lineage, identity as cholinergic neurons, connectivity and provide insight into their function during mating. (see poster by R.C. Bonnington). PHD specification is unaffected in
cdk-4 mutants and the majority of nom mutants tested. This suggests that largely distinct genetic mechanisms regulate the two events. The male provides two glia-to-neuron cell fate switches that occur by seemingly independent cellular and genetic mechanisms. We are continuing to investigate the regulatory strategies that confer neurogenic potential to differentiated glia.