野村 隆士

カベオラがエンドサイトーシスを行う膜ドメインであるかどうかは，未だに議論されている問題である．しかし，Simian virus 40（SV40）の細胞内侵入のリアルタイムイメージング解析により，ウイルス侵入に利用されるカベオラエンドサイトーシスの詳細が明らかになってきた．SV40を内包したカベオラは，ダイナミン依存的に細胞膜からbuddingする．その後，SV40はカベオソームへ移動し，やがてERへと運ばれる．これに加えて，ヒトコロナウイルス-229Eも細胞内侵入にカベオラを利用することがわかってきた．本稿では，現在明らかにつつあるカベオラエンドサイトーシスを紹介し，ウイルス侵入との関わり合いを論じたいと思う．

The adenomatous polyposis coli (APC) tumor suppressor gene is linked to the development of familial adenomatous polyposis and sporadic colorectal tumors. Though APC is highly expressed in the central nervous system (CNS) and directly binds to PSD-95 through the extreme C terminus of APC, the significance of the interaction between APC and PSD-95 in CNS remains unknown. In this study, we investigated whether APC associates neural signal transduction via clustering of PSD-95 in neurons. We performed the calcium imaging and the patch clamp recording in 9-11 days cultured hippocampal neurons. Bath application of 1 μM AMPA increased intracellular calcium concentration in control cells. However, this response was inhibited in the dominant negative (APC-C72) cells, which were overexpressed a dominant negative molecule (APC-C72) that could disrupt the interaction between APC and PSD-95. Under patch clamp condition, no interactions were observed in the neighboring APC-C72 cells. These results suggest that APC may affect AMPA signal transduction via clustering of PSD-95. <b>[Jpn J Physiol 54 Suppl:S147 (2004)]</b>

Caveolae and rafts are two different domains of the plasma membrane. Caveolae are small invaginations and characterized by the presence of unique membrane proteins named caveolins. Caveolins bind cholesterol, form oligomers, interact with various signaling molecules, and induce caveolae formation. On the other hand, rafts are highly dynamic structure and do not take any characteristic shape; but both caveolae and rafts are enriched with sphingolipids and cholesterol and appear to contain many molecules in common. Some of the functions which have been ascribed to caveolae, such as signal transduction, may be mediated by both caveolae and rafts. Caveolins are thought to be involved in the intracellular cholesterol transport, and an increase or decrease of cholesterol content appears to affect caveolae and rafts. Mutation of caveolin genes or quantitational change of caveolae and caveolins has been reported to occur in several diseases. Further studies should elucidate the physiological roles as well as pathological changes of these functional domains in the plasma membrane.

Calreticulin (CRT)-1 is a major Ca2+-buffering protein in the lumen of the endoplasmic reticulum. Human and murine CRT-2 was isolated in 2002, but the subcellular localization and function is still unclear. Here, we studied the intracellular localization and function of CRT-2 with hemagglutinin-tagged (HA-) human CRT-2. Western blotting revealed HA-CRT-2 as a single band at 50 kDa. Using immunofluorescence microscopy of cultured fibroblasts and epithelial cells transfected with HA-CRT-2 cDNA, labeling for HA-CRT-2 was seen as a reticular network with a nuclear envelope pattern that colocalized with calnexin and protein disulfide isomerase. Immunoelectron microscopy confirmed that HA-CRT-2 was localized in the lumen of the endoplasmic reticulum. Stains-all staining, a method to detect Ca2+-binding proteins, could not stain the immunoprecipitate of HA-CRT-2, although HA-CRT-1 immunoprecipitate was stained blue. These results indicate that the molecular weight of the non-tagged CRT-2 on SDS-PAGE is 49 kDa, and that CRT-2, as well as CRT-1, is localized in the lumen of the endoplasmic reticulum, but that CRT-2 capacity for Ca2+-binding may be absent or much lower than that of CRT-1. ©