Abstract
Rac GTPases act as master switches to coordinate multiple interweaved signaling pathways. A major function for Rac GTPases is to control neurite development by influencing downstream effector molecules and pathways. In Caenorhabditis elegans, the Rac proteins CED-10, RAC-2 and MIG-2 act in parallel to control axon outgrowth and guidance. Here, we have identified a single glycine residue in the CED-10/Rac1 Switch 1 region that confers a non-redundant function in axon outgrowth but not guidance. Mutation of this glycine to glutamic acid (G30E) reduces GTP binding and inhibits axon outgrowth but does not affect other canonical CED-10 functions. This demonstrates previously unappreciated domain-specific functions within the CED-10 protein. Further, we reveal that when CED-10 function is diminished, the adaptor protein NAB-1 (Neurabin) and its interacting partner SYD-1 (Rho-GAP-like protein) can act as inhibitors of axon outgrowth. Together, we reveal that specific domains and residues within Rac GTPases can confer context-dependent functions during animal development.
Highlights
Correct axonal projection is essential for developing a functional nervous system
Previous studies have shown that molecular switch proteins called Rac GTPases perform redundant functions in controlling neurite development
Using a pair of bilateral neurons in the nematode Caenorhabditis elegans to model neurite development, we found that a single amino acid in a conserved domain of the Rac GTPase CED-10 is crucial for controlling neurite outgrowth in a partially non-redundant manner
Summary
All C. elegans strains were maintained at 20 ̊C on NGM plates seeded with Escherichia coli OP50 bacteria, unless otherwise stated [65]. Strains were generated using standard genetic procedures and are listed in S3 and S4 Tables. All strains were backcrossed to N2 at least three times before scoring or generating compound mutants. Genotypes were confirmed using PCR genotyping or Sanger sequencing with primers listed in S5 Table. Neurons were scored, blinded to phenotype, in L4/young adult stages, unless otherwise stated. Scoring criteria for each neuron is detailed in S2 Table. The following transgenes were used to enable visualization of specific neurons. All neuronal scoring was repeated in triplicates on independent days, n = 75, except for oxIs12 scoring, n = 35
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