The Cloning and Characterization of MADD-4, A Novel Guidance Cue in C. elegans

Directed cell migration is fundamental to the development of all multicellular organisms including humans. To investigate the molecular underpinnings of directed cell migration, our lab primarily exploits plasma membrane extensions called muscle arms, from the body wall muscles in the tiny nematode Caenorhabditis elegans. Through genetic screens for genes required for muscle arm extension, we found that the UNC-40/DCC netrin guidance receptor directs muscle arm extension to the midlines. Surprisingly, neither the UNC-6/netrin cue (the canonical ligand for UNC-40) nor other well-characterized guidance cues such as the slits, ephrins and semaphorins were found to be the primary cue for muscle arm extension. This suggested that muscle arms were likely responding to a novel guidance cue. In this thesis, I describe the cloning and characterization of MADD-4, a novel secreted cue that diffuses and attracts muscle arms and sensory axons along the dorsoventral axis in C. elegans. MADD-4 is a member of the non-enzymatic ADAMTSL family of proteins and is well conserved among animals. Very little is known about the biological role of any of MADD-4's orthologs. Together with Kevin Chan, I found that MADD-4's guidance function is dependent on an EVA-1-UNC-40 co-receptor complex. We found that MADD-4 interacts with both EVA-1 and UNC-40. Similarly, we found that EVA-1 and UNC-40 likely physically interact and this interaction is critical for the MADD-4 response. Furthermore, we found that the binding of EVA-1 to UNC-40 increases UNC-40's sensitivity to MADD-4. This enhanced sensitivity becomes especially meaningful within a field of other ligands capable of binding UNC-40 like UNC-6. In the absence of UNC-6, UNC-40's responsiveness to MADD-4 becomes less dependent on EVA-1. Hence, by regulating UNC-40's sensitivity to MADD-4, EVA-1 may increase the precision by which UNC-40-directed processes can reach MADD-4-expressing target cells. Collectively, the work discussed in this thesis recounts the first description of a novel guidance cue and its mechanism of action. Furthermore, since the biological role of any ADAMTSL family member outside of MADD-4 is largely unknown, it is very likely that my work on MADD-4 will broaden our understanding of the biological role of the ADAMTSL family of proteins.Ph.D.2018-06-17 00:00:0