Kallman Syndrome (KS) is a human genetic disorder resulting from aberrant olfactory axon guidance. Two proteins have been implicated in KS, the novel secreted protein anosmin-1 and FGFR 1. The signaling pathways regulated by anosmin-1 have remained unclear, and in vivo evidence linking anosmin-1 to FGF signaling is still lacking. We are using C. elegans to elucidate these questions. The C. elegans ortholog of anosmin-1, KAL-1, has been implicated in embryonic morphogenesis and axon branching.
kal-1 null mutants display low penetrance epidermal morphogenetic defects. Using 4-D microscopy and quantitative analysis of neuroblast (NB) migration timing, we find that the primary defect in
kal-1 mutants is in migration of ventral neuroblasts during gastrulation cleft closure. Double mutant analysis indicates that KAL-1 likely acts in parallel to axon guidance pathways (VAB-1/EphR, MAB-20/Semaphorin, EFN-4).Genetic evidence from C.elegans suggests that KAL-1 signaling may require heparan sulfated proteoglycans (HSPGs). Mutations in genes encoding HS modification enzymes C-5 epimerase (
hse-5) and 6-O-sulfotransferase (
hst-6) suppress ectopic axon branching caused by overexpression of KAL-1. We find that null alleles of
hse-5 and
hst-6 phenocopy
kal-1 NB migration defects and do not enhance
kal-1 phenotypes, suggesting that KAL-1 signaling in the embryo, as in axon branching, involves interaction with HSPGs modified by these enzymes. Mutations in other sulfotransferases (
hst-2,
hst-3) do not affect NB migration. Which HSPGs are involved in KAL-1 signaling? Loss of function of the HSPG core proteins syndecan (
sdn-1) and glypican (
gpn-1) causes weak NB migration defects. However
gpn-1 sdn-1 double mutants resemble
kal-1 in NB migration, suggesting that these two HSPGs have redundant roles in NB migration. In preliminary biochemical experiments a KAL-1 affinity matrix selectively bound GPN-1 and SDN-1 from extracts of C. elegans embryos. Thus, genetic and biochemical evidence suggests KAL-1 may interact redundantly with either GPN-1 or SDN-1. KAL-1 appears to be expressed on the surfaces of ventral neuroblasts and may act as a short range cue in cell adhesion or repulsion. We are determining where GPN-1 and SDN-1 function is required in NB migration. We will also present our analysis of whether FGF signaling is involved in KAL-1 mediated processes in the embryo.