Duration of fusion pore opening and the amount of hormone released are regulated by myosin II during kiss-and-run exocytosis

Ryo Aoki, Tetsuya Kitaguchi, Manami Oya, Yu Yanagihara, Mai Sato, Atsushi Miyawaki, Takashi Tsuboi

Research output: Contribution to journalArticle

29 Citations (Scopus)

Abstract

Since the fusion pore of the secretory vesicle is resealed before complete dilation during 'kiss-and-run' exocytosis, their cargoes are not completely released. Although the transient fusion pore is kept open for several seconds, the precise mechanisms that control fusion pore maintenance, and their physiological significance, are not well understood. Using dual-colour TIRF (total internal reflection fluorescence) microscopy in neuroendocrine PC12 cells, we show that myosin II regulates the fusion pore dynamics during kiss-and-run exocytosis. The release kinetics of mRFP (monomeric red fluorescent protein)-tagged tPA (tissue plasminogen activator) and Venus-tagged BDNF (brain-derived neurotrophic factor), which show slower release kinetics than NPY (neuropeptide Y)-mRFP and insulin-mRFP, were prolonged by the overexpression of a wild-type form of the RLC (myosin II regulatory light chain). In contrast, overexpression of a dominant-negative form of RLC shortened the release kinetics. Using spH (synapto-pHluorin), a green fluorescent protein-based pH sensor inside the vesicles, we confirmed that themodulation of the release kinetics by myosin II is due to changes in the duration of fusion pore opening. In addition, we revealed that the amount of hormone released into the extracellular space upon stimulation was increased by overexpression of wild-type RLC. We propose that the duration of fusion pore opening is regulated by myosin II to control the amount of hormone released from a single vesicle.

Original languageEnglish
Pages (from-to)497-504
Number of pages8
JournalBiochemical Journal
Volume429
Issue number3
DOIs
Publication statusPublished - 2010 Aug 1
Externally publishedYes

Fingerprint

Myosin Type II
Exocytosis
Myosin Light Chains
Fusion reactions
Hormones
Kinetics
Venus
Neuroendocrine Cells
Neuropeptide Y
PC12 Cells
Brain-Derived Neurotrophic Factor
Secretory Vesicles
Extracellular Space
Tissue Plasminogen Activator
Green Fluorescent Proteins
Fluorescence Microscopy
pH sensors
Dilatation
Color
Fluorescence microscopy

Keywords

  • Actin
  • Dense-core vesicle
  • Exocytosis
  • Myosin II
  • Total internal reflection fluorescence microscopy (TIRF microscopy)

ASJC Scopus subject areas

  • Biochemistry
  • Cell Biology
  • Molecular Biology
  • Medicine(all)

Cite this

Duration of fusion pore opening and the amount of hormone released are regulated by myosin II during kiss-and-run exocytosis. / Aoki, Ryo; Kitaguchi, Tetsuya; Oya, Manami; Yanagihara, Yu; Sato, Mai; Miyawaki, Atsushi; Tsuboi, Takashi.

In: Biochemical Journal, Vol. 429, No. 3, 01.08.2010, p. 497-504.

Research output: Contribution to journalArticle

Aoki, R, Kitaguchi, T, Oya, M, Yanagihara, Y, Sato, M, Miyawaki, A & Tsuboi, T 2010, 'Duration of fusion pore opening and the amount of hormone released are regulated by myosin II during kiss-and-run exocytosis', Biochemical Journal, vol. 429, no. 3, pp. 497-504. https://doi.org/10.1042/BJ20091839
Aoki, Ryo ; Kitaguchi, Tetsuya ; Oya, Manami ; Yanagihara, Yu ; Sato, Mai ; Miyawaki, Atsushi ; Tsuboi, Takashi. / Duration of fusion pore opening and the amount of hormone released are regulated by myosin II during kiss-and-run exocytosis. In: Biochemical Journal. 2010 ; Vol. 429, No. 3. pp. 497-504.
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