Myosin va mediates docking of secretory granules at the plasma membrane.
Myosin Va (MyoVa) is a prime candidate for controlling actin-based organelle motion in neurons and neuroendocrine cells. Its function in secretory granule (SG) trafficking was investigated in enterochromaffin cells by wide-field and total internal reflection fluorescence microscopy. The distribution of endogenous MyoVa partially overlapped with SGs and microtubules. Impairing MyoVa function by means of a truncated construct (MyoVa tail) or RNA interference prevented the formation of SG-rich regions at the cell periphery and reduced SG density in the subplasmalemmal region. Individual SG trajectories were tracked to analyze SG mobility. A wide distribution of their diffusion coefficient, D(xy), was observed. Almost immobile SGs (D(xy) < 5 x 10(-4) microm2 x s(-1)) were considered as docked at the plasma membrane based on two properties: (1) SGs that undergo exocytosis have a D(xy) below this threshold value for at least 2 s before fusion; (2) a negative autocorrelation of the vertical motion was found in subtrajectories with a D(xy) below the threshold. Using this criterion of docking, we found that the main effect of MyoVa inhibition was to reduce the number of docked granules, leading to reduced secretory responses. Surprisingly, this reduction was not attributable to a decreased transport of SGs toward release sites. In contrast, MyoVa silencing reduced the occurrence of long-lasting, but not short-lasting, docking periods. We thus propose that, despite its known motor activity, MyoVa directly mediates stable attachment of SGs at the plasma membrane.
Desnos
Claire
C
Institut de Biologie Physico-Chimique, Centre National de la Recherche Scientifique, Unité Propre de Recherche 1929, Université Paris 7 Denis Diderot, 75005 Paris, France.
Huet
Sébastien
S
Fanget
Isabelle
I
Chapuis
Catherine
C
Böttiger
Caroline
C
Racine
Victor
V
Sibarita
Jean-Baptiste
JB
Henry
Jean-Pierre
JP
Darchen
François
F
D002462:Cell Membrane / Q000502:physiology*; D002478:Cells, Cultured; D006801:Humans; D018995:Myosin Heavy Chains / Q000502:physiology*; D024701:Myosin Type V / Q000502:physiology*; D022142:Secretory Vesicles / Q000502:physiology*; D022161:Transport Vesicles / Q000502:physiology
C077511:MYO5A protein, human; D024701:Myosin Type V; D018995:Myosin Heavy Chains