Reduce, Reuse, Recycle: The tale of two Wnts and the lone C. elegans Syndecan, SDN-1

Abstract

Heparan sulfate proteoglycans (HSPGs) are cell adhesion molecules that have been shown to be involved in a myriad of different aspects of development such as embryogenesis, dorsal-ventral axon guidance and cell migration. Despite knowing the phenotypes caused by mutations in HSPGs, little is known about the ligands working with HSPGs during development. Identification of HSPG ligands will provide insight into how HSGPs function during embryogenesis and later in neural development. Chapter II describes two Caenorhabditis elegans cell adhesion proteins SDN-1/Syndecan and PTP-3/LAR-RPTP as important regulators of polarization and cell migration during embryogenesis. Loss-of-function (LOF) mutations in either ptp-3 or sdn-1 resulted in low penetrance embryonic developmental defects. We used double mutant analysis to test whether ptp-3 and sdn-1 function in a linear genetic pathway during C. elegans embryogenesis and found that double mutants of sdn-1 and ptp-3 exhibited a highly penetrant synthetic lethality (SynLet), with only a small percentage of animals surviving to adulthood. Analysis of the survivors demonstrated that these animals had a synergistic increase in the penetrance of embryonic developmental defects. Taken together, these data strongly suggested that PTP-3 and SDN-1 function in parallel during embryogenesis. We subsequently used RNAi to knockdown the function of ~3,600 genes predicted to encode secreted and/or transmembrane molecules to identify genes that interacted with ptp-3 or sdn-1. We found that the Wnt ligand, lin-44, was SynLet with sdn-1 but not ptp-3. We used 4-dimensional time-lapse analysis to characterize the interaction between lin-44 and sdn-1. We found evidence that loss of lin-44 caused defects in the polarization and migration of endodermal precursors during gastrulation, a previously undescribed role for lin-44 that is strongly enhanced by the loss of sdn-1. In chapter III the interaction between SDN-1, LIN-44 and EGL-20 during DD/VD (Dorsal D-type and Ventral D-type motorneurons, respectively) axon outgrowth and termination is described. Double mutant analysis between sdn-1, lin-44 and egl-20 suggests SDN-1 acts extrinsically to inhibit the activation of BAR-1/β-catenin during axon outgrowth of the D-type motorneurons specifically, DD6 and VD13. Then SDN-1 acts intrinsically within the Wnt signaling pathway to ensure proper outgrowth termination of those axons. These results show for the first time that the same Wnt signaling pathways are both positively and negatively regulated by SDN-1 during axon growth. Altogether, our genetic analysis suggest that axon outgrowth and termination occurs in two steps with Wnt ligands acting with SDN-1 in a combinatorial fashion and are not simply working in a parallel manner as previously reported.