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A small GTPase, human Rab32, is required for the formation of autophagic vacuoles under basal conditions

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Abstract

Here we show that a small GTPase, Rab32, is a novel protein required for the formation of autophagic vacuoles. We found that the wild-type or GTP-bound form of human Rab32 expressed in HeLa and COS cells is predominantly localized to the endoplasmic reticulum (ER), and overexpression induces the formation of autophagic vacuoles containing an autophagosome marker protein LC3, the ER-resident protein calnexin and endosomal/lysosomal membrane protein LAMP-2, even under nutrient-rich conditions. The recruitment of Rab32 to the ER membrane was necessary for autophagic vacuole formation, suggesting involvement of the ER as a source of autophagosome membranes. In contrast, the expression of the inactive form of, or siRNA-specific for, Rab32 caused the formation of p62/SQSTM1 and ubiquitinated protein-accumulating aggresome-like structures and significantly prevented constitutive autophagy. We postulate that Rab32 facilitates the formation of autophagic vacuoles whose membranes are derived from the ER and regulates the clearance of aggregated proteins by autophagy.

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Acknowledgments

We thank Drs. Ikuo Wada (Fukushima Medical University) and Tomoki Chiba (University of Tsukuba) for providing the ER-cherry and FLAG-tagged ubiquitin constructs, respectively. This study was supported in part by a grant-in-aid from the Ministry of Education, Culture, Sports, Sciences and Technology of Japan. Yuko Hirota is a Research Fellow of the Japan Society for the Promotion of Science.

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  1. Division of Pharmaceutical Cell Biology, Graduate School of Pharmaceutical Sciences, Kyushu University, 3-1-1 Maidashi, Fukuoka, 812-8582, Japan

    Yuko Hirota & Yoshitaka Tanaka

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  1. Yuko Hirota
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Correspondence to Yoshitaka Tanaka.

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Hirota, Y., Tanaka, Y. A small GTPase, human Rab32, is required for the formation of autophagic vacuoles under basal conditions. Cell. Mol. Life Sci. 66, 2913–2932 (2009). https://doi.org/10.1007/s00018-009-0080-9

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