RPM-1, a Caenorhabditis elegans protein that functions in presynaptic differentiation, negatively regulates axon outgrowth by controlling SAX-3/robo and UNC-5/UNC5 activity

Changes in axon outgrowth patterns are often associated with synaptogenesis. Members of the conserved Pam/ Highwire/ RPM- 1 protein family have essential functions in presynaptic differentiation. Here, we show that Caenorhabditis elegans RPM- 1 negatively regulates axon outgrowth mediated by the guidance receptors SAX- 3/ robo and UNC- 5/ UNC5. Loss- of- function rpm- 1 mutations cause a failure to terminate axon outgrowth, resulting in an overextension of the longitudinal PLM axon. We observe that PLM overextension in rpm- 1 mutants is suppressed by sax- 3 and unc- 5 loss- of- function mutations. PLM axon overextension is also induced by SAX- 3 overexpression, and the length of extension is enhanced by loss of rpm- 1 function or suppressed by loss of unc- 5 function. We also observe that loss of rpm- 1 function in genetic backgrounds sensitized for guidance defects disrupts ventral AVM axon guidance in a SAX- 3- dependent manner and enhances dorsal guidance of DA and DB motor axons in an UNC- 5- dependent manner. Loss of rpm- 1 function alters expression of the green fluorescent protein ( GFP)- tagged proteins, SAX- 3:: GFP and UNC- 5:: GFP. RPM- 1 is known to regulate axon termination through two parallel genetic pathways; one involves the Rab GEF ( guanine nucleotide exchange factor) GLO- 4, which regulates vesicular trafficking, and another that involves the F- box protein FSN- 1, which mediates RPM- 1 ubiquitin ligase activity. We show that glo- 4 but not fsn- 1 mutations affect axon guidance in a manner similar to loss of rpm- 1 function. Together, the results suggest that RPM- 1 regulates axon outgrowth affecting axon guidance and termination by controlling the trafficking of the UNC- 5 and SAX- 3 receptors to cell membranes.