The Ret receptor mediates sensory neuron dendrite growth through TGFβ

The development of a functional nervous system requires correct neuronal specification to allow proper sensory information processing and communication within the neuronal network. In particular, neuronal dendrite development is highly complex and therefore strictly regulated by a variety of substrate and tissue interaction. Neurons establish subtype specific, often highly stereotyped branching patterns to cover their receptive field. Cell type specific functional complexity correlates with the morphological complexity characterized by the dendritic shape, arborization and size (Ramon y Cajal S, 1911). However, due to this complexity, mechanistic and molecular insight into dendrite development is still limited. The Drosophila melanogaster larval peripheral nervous system (PNS) is a powerful genetic model for the analysis of dendrite development and function. Within the larval PNS, the well characterized dendritic arborization (da) neurons provide an excellent system to study dendrite development in vivo. C4da neurons feature class specific highly complex dendritic fields tiling the entire larval body wall. In this study, the conserved receptor tyrosine kinase (RTK) Ret (re-arranged during transfection) has been identified as an important regulator of C4da neuron dendrite development and patterning. A newly generated Ret knock-out (Retko) allele revealed strong C4da neuron dendrite growth and adhesion defects. This phenotype could be fully rescued by C4da neuron specific expression of transgenic Ret in Retko animals. Subsequent structure-function analysis of Ret revealed that the extra- and intracellular domain are both required for C4da dendrite development indicating that Ret transduces extracellular signals into the cell. Surprisingly, Ret tyrosine kinase activity is dispensable for Ret function in dendrite development, as expression of kinase-inactive Ret showed complete rescue activity. These results provide evidence that Ret is cell-autonomously required for C4da neuron dendrite patterning independently of its kinase activity. Furthermore, Retko animals displayed behavioural defects linked to C4da neuron function. A candidate screen for Ret ligands identified the TGFβ ligand maverick (mav) as a potential interactor mediating Ret function in C4da neuron development. Mav is likely expressed and secreted by epithelial cells and analysis of mav function by generation of a mav knock-out allele (mavko) confirmed a role for mav in Ret dependent C4da neuron development. Moreover, mav likely acts as a permissive guidance signal required for C4da neuron dendrite patterning. Additionally, overexpression of mav and Ret is sufficient to induce dendrite growth in heterologous neurons suggesting that mav/Ret signaling is able to transduce growth signals. Overall, the data provide evidence that Ret and mav are acting in same signaling pathway in C4da neuron development, potentially as a novel receptor-ligand protein complex. Analysis of the underlying signaling pathway indicated that canonical TGFβ signaling is not involved in Ret and mav function. Instead, genome wide microarray profiling of Retko C4da neurons identified RanBPM as a potential mediator of Ret intracellular signaling. Further analysis showed that RanBPM is expressed in C4da neurons and its loss-of-function displays Ret-like phenotypes in C4da neurons. In addition, a functional genetic link between RanBPM and Ret in C4da neurons development could be identified suggesting that RanBPM is a novel component of the Ret signaling pathway. This work provides novel insight into the Ret signaling pathway required for C4da neuron dendrite development, including the identification of mav as a potential ligand. Based on the high degree of conservation of Ret, these findings suggest potentially conserved functions in dendrite development of vertebrates.