| Summary: | 碩士 === 長庚大學 === 基礎醫學研究所 === 90 === Abstract
Using both forward and reverse genetics in Caenorhabditis elegans several proteins involved in yolk transport were recently identified. Among them, RME-2 has been proved to be the yolk receptor. It associates primarily with small vesicles immediately below the plasma membrane of oocytes. RME-1, which has an N-terminal P-loop nucleotide-binding domain, a central coil-coil domain, and a C-terminal Eps15 homology (EH) domain, was implied in the control of exit of recycling endosomes in mammalian cells. In this study we dissect the molecular mechanism of RME-1 by examining dominant negative rme-1 mutants with immuno-electron microscopy. We demonstrated that defects in molecular structure of RME-1 resulted in shifting of RME-2 population from a submembrane locale to multivesicular structures deeper within the oocytes. Therefore, the RME-1 functions in yolk receptor recycling can be inferred. In the intestine, the RME-1 molecules were localized to basolateral plasmalemma and tubulo-vesicular membrane sectors, the lumen of which became dilated upon an amino acid change of RME-1 molecule near the EH-domain. In both cases, the labeling of RME-1 could be seen at the neck region of vesicles invaginating from plasma membrane or pinching off from the tubulo-vesicular structures. On the other hand, mutation affecting the P-loop resulted in diffuse RME-1 labeling pattern. Taken together, this indicates the importance of P-loop domain in proper endosomal targeting and EH domain in vesicle budding.
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