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Role of VAMP-2, VAMP-7, and VAMP-8 in constitutive exocytosis from HSY cells

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Abstract

We evaluated the role of VAMP-2/synaptobrevin, VAMP-7/TI-VAMP, and VAMP-8/endobrevin in exocytic pathways of HSY cells, a human parotid epithelial cell line, by coexpressing these VAMP proteins tagged with green fluorescent protein (GFP) and human growth hormone (hGH) as a secretory cargo. Exocytosis of hGH was constitutive and the fluorescent signal of hGH–GFP was observed in the Golgi area and small vesicles quickly moving throughout the cytoplasm. The cytoplasmic vesicles containing hGH overlapped well with VAMP-7-GFP, but did so scarcely with VAMP-2-GFP or VAMP-8-GFP. However, when the vesicle transport from the trans-Golgi network to the plasma membrane was arrested by incubation at 20°C for 2 h and then released by warming up to 37°C; VAMP-2-GFP and hGH were clearly colocalized together in small cytoplasmic vesicles. Neither VAMP-7-GFP nor hGH–GFP was colocalized with LAMP-1, a marker for lysosomes and late endosomes. These results suggest that (1) VAMP-2 can be one of the v-SNAREs for constitutive exocytosis; (2) VAMP-7 is involved in the constitutive exocytosis as a slow, minor v-SNARE, but not in the lysosomal transport; and (3) VAMP-8 is unlikely to be a v-SNARE for constitutive exocytosis in HSY cells.

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Abbreviations

GFP:

Green fluorescent protein

SNARE:

SNAP receptor

VAMPs:

Vesicle-associated membrane proteins

TI-VAMP:

Tetanus toxin-insensitive VAMP

hGH:

Human growth hormone

LAMP-1:

Lysosome-associated membrane protein 1

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Acknowledgements

We wish to thank Prof. Yosuke Tojyo, as well as Drs. Takao Morita and Akihiko Nezu, Department of Dental Pharmacology, Health Sciences University, for helpful discussion and also Ms. Tamaki Tamaki-Yokohama and Mr. Kouhei Taima for their technical assistance. This work was supported in part by the Academic Sciences Frontier Project and by a grant-in-aid for scientific research (No.15591973 to T. T.) from the Ministry of Education, Culture, Sports, Science, and Technology of Japan.

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Authors and Affiliations

  1. Department of Oral Biochemistry, School of Dentistry, Health Sciences University of Hokkaido, Tobetsu, Hokkaido, 061-0293, Japan

    Yohei Oishi, Toshiya Arakawa & Taishin Takuma

  2. Department of Orthodontics, School of Dentistry, Health Sciences University of Hokkaido, Tobetsu, Hokkaido, 061-0293, Japan

    Yohei Oishi & Itaru Mizoguchi

  3. Department of Dental Pharmacology, School of Dentistry, Health Sciences University of Hokkaido, Tobetsu, Hokkaido, 061-0293, Japan

    Akihiko Tanimura

  4. Department of Biochemistry, Kitasato University School of Medicine, Sagamihara, Kanagawa, 228-8555, Japan

    Makoto Itakura & Masami Takahashi

  5. Department of Oral Pathology, School of Dentistry, Meikai University, Sakado, Saitama, 350-0283, Japan

    Yoshifumi Tajima

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  1. Yohei Oishi
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Correspondence to Taishin Takuma.

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Oishi, Y., Arakawa, T., Tanimura, A. et al. Role of VAMP-2, VAMP-7, and VAMP-8 in constitutive exocytosis from HSY cells. Histochem Cell Biol 125, 273–281 (2006). https://doi.org/10.1007/s00418-005-0068-y

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  • DOI: https://doi.org/10.1007/s00418-005-0068-y

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