Affiliations: 1) Program of Biomedical Science, Graduate School of Integrated Sciences for Life, Hiroshima University, 1-3-1 Kagamiyama, Higashi-Hiroshima, Hiroshima 739-8526, Japan; 2) Program of Basic Biology, Graduate School of Integrated Sciences for Life, Hiroshima University, 1-3-1 Kagamiyama, Higashi-Hiroshima, Hiroshima 739-8526, Japan
Keywords: a. neural degeneration; e. neuronal morphogenesis
VAP is a type II integral transmembrane protein localized at the endoplasmic reticulum (ER), and functions as a tethering protein of the membrane contact sites between ER and various intracellular organelles. Recent studies revealed that VAP is cleaved, and its N-terminal MSP domain is secreted to the extracellular space in C. elegans, Drosophila and also in human. The secreted VAP MSP domain is known to bind to a variety of axon growth cone guidance receptors (Eph, Robo, Lar etc.), implying that VAP could function non-cell autonomously. In addition, mutations in human VAPB were known to cause neurodegenerative diseases such as amyotrophic lateral sclerosis 8 (ALS8). However, as the VAP loss-of-function exhibits severe lethality in early developmental stages, the physiological functions of VAP, especially the extracellular functions of VAP MSP domain are not well understood.
In order to investigate the physiological functions of VAP (Drosophila ortholog is Vap33), we utilized Drosophila olfactory projection neurons with a mosaic analysis with a repressible cell marker (MARCM), which allows us to analyze the function of a gene of interest in a single cell resolution in vivo. In vap33-/- MARCM clones, the dendrite but not axon of projection neurons exhibited the severe morphological defects, which are rescued by the cell-autonomous expression of vap33 cDNA, suggesting that VAP cell-autonomously and preferentially regulates dendrite morphology. Interestingly, subcellular localizations of Golgi apparatus and mitochondria were also severely affected. These results indicate the requirement of intracellular functions of Vap33 in neural development. Moreover, to examine the roles of the secreted Vap33 MSP domain, we searched the important sequence required for Vap33 MSP secretion and succeeded to generate the “unsecretable vap33 mutant” whose Vap33 MSP is not secreted but can function intracellularly. The unsecretable vap33 mutant exhibited lethality. Overexpressing the unsecretable Vap33 in the vap33-/- PN clones rescued the dendritic defects, implying that the intracellular functions of unsecretable Vap33 are likely to be normal. From these results, we propose that secreted MSP domain plays important extracellular functions to maintain organismal viability. We’d like to discuss how secreted MSP domain contributes the animal physiology, which may lead the clues for the therapeutic strategy of neurodegenerative diseases.