The correct outgrowth of axons is crucial for the establishment of neuronal networks during embryonic development. To identify novel genes involved in axon guidance and fasciculation we performed a direct visual screen for mutants with outgrowth defects in
glr-1(glutamate receptor)::GFP expressing interneurons. Two alleles of
ast-1(axonal steereing) called
rh300 and
hd1 were recovered independently from these screens. Their phenotypes are characterized by defasciculated interneuron axons in the ventral cord, where axons cross from the right to the left bundle. Occasionally individual interneuron axons do not reach the ventral cord but run in a lateral position instead. The penetrance of these defects combined is about 40%. The mutants are not significantly uncoordinated despite these defects in the command interneurons of the motorcircuit. There are no obvious defects in the navigation of other axons. Non-neuronal tissues and cell migration appear unaffected in
ast-1 mutant animals. Rescue of the mutant phenotype was obtained by injection of cosmid H37P12 and with a PCR fragment containing the predicted open reading frame T08H4.3. This gene encodes a transcription factor with an ETS DNA-binding domain. Sequencing the DNA of the two mutant alleles revealed point mutations in exon six causing amino acid exchanges in the highly conserved region of the ETS domain. ETS proteins are targets of the Ras-MAPK signaling pathway and interact with other transcription factors that promote the binding of ETS proteins to composite Ras responsive elements. The C. elegans genome contains ten ETS genes. Ast-1 and C42D8.4 are closely related to the mammalian
fli-1 subfamily. The ventral cord midline crossing defects characteristic for
ast-1 mutants are also seen in several other mutants in genes controlling aspects of neuronal differentiation. Using double mutant analysis we found that
ast-1 genetically interacts with the transcription factors
zag-1 and
lin-11. In addition, AST-1 shows synergistic effects with the netrin/UNC-6 receptor UNC-40 and the actin-regulating protein UNC-34. Currently we are trying to further elucidate the function of
ast-1 in the regulatory network of axon guidance genes.