| Proper functioning of the nervous system relies heavily on the precise, yet complex connections between axons and their target. In both vertebrates and invertebrates, the limited numbers of axon guidance cues and their receptors play a pivotal role in achieving this task. The main goal of my study is to elucidate mechanisms by which a limited set of guidance cues might orchestrate such diverse morphological outcomes.;The Drosophila transmembrane semaphorin Sema-la is a repulsive guidance cue that utilizes the Plexin A (PlexA) receptor during neural development. Sema-la is required in axons to facilitate axon defasciculation at guidance choice points during embryonic motor axon guidance. To identify novel modulatory components of the Sema-la signaling pathway, I embarked on an unbiased genetic modifier screen on the X chromosome utilizing the Sema-la gain of function (GOF) phenotype in the CNS axonal scaffold, and identified several candidate genes.;Among several candidates, I identified the Trol gene, mutations in which strongly suppress Sema-la---mediated repulsive axon guidance. Trol encodes the phylogenetically conserved secreted heparan sulfate proteoglycan (HSPG) perlecan, a component of the extracellular matrix. Motor axon guidance defects in perlecan mutants resemble those observed in Sema-la and PlexA mutant embryos, and perlecan mutants genetically interact with PlexA and Sema-la mutants. Perlecan protein is found both in the CNS and in the peripheral nervous system, with higher expression levels in the vicinity of motor axon trajectories and pathway choice points. Restoring perlecan to mutant motor neurons rescues perlecan axon guidance defects. Perlecan augments the reduction in phospho-focal adhesion kinase (FAK) levels that result from treating insect cells in vitro with Sema-la, and genetic interactions among integrin, Sema-la , and FAK in vivo support an antagonistic relationship between Sema-la and integrin signaling. Taken together, these observations show that perlecan is required for Sema-la---mediated motor axon guidance, revealing roles for extracellular matrix proteoglycans in modulating transmembrane guidance cue signaling during neural development in vivo.;The knowledge of extracellular matrix proteoglycan perlecan facilitating repulsive transmembrane sempahorin signaling provides insight into how the surrounding environment of growing axons can modulate guidance cue signaling. It is likely that similar mechanisms exist in the vertebrate nervous system during development or in the adult at sites of nervous system injury, thus my study may contribute to the development of approaches directed toward inducing functional recovery following axon regeneration after injury. |
