Multiple Mechanisms Driving F-actin-Dependent Transport of Organelles to and From Secretory Sites in Bovine Chromaffin Cells

Abstract

Neuroendocrine chromaffin cells represent an excellent model to study the molecular mechanisms associated with the exo-endocytotic cycle of neurotransmitter release. In this study, EGFP-Lifeact and confocal microscopy has been used to analyze the re-organization of the cortical F-actin cytoskeleton associated to organelle transport during secretion with unprecedented detail. In these cells secretory events accumulate in temperature-sensitive and myosin II-dependent F-actin expansions and retractions affecting specific regions of the sub-membrane space. Interestingly, not only vesicles but also mitochondria are transported toward the plasmalemma during these expansions. Simultaneously, we found F-actin cytoskeletal retraction withdraws vesicles from the sub-plasmalemmal space, forming novel empty internal spaces into which organelles can be transported. In addition to these well-coordinated, F-actin-myosin II dependent processes that drive the transport of the majority of vesicles, fast transport of chromaffin vesicles was observed, albeit less frequently, which used F-actin comet tails nucleated from the granular membrane. Thus, upon cell stimulation F-actin structures use diverse mechanisms to transport organelles to and from the membrane during the exo-endocytotic cycle taking place in specific areas of cell periphery.

MUNC18-1 regulates the submembrane F-actin network, independently of syntaxin1 targeting, via hydrophobicity in β-sheet 10

“…In the cortical regions underneath the PM of chromaffin cells, a dense F-actin network is the main component of the cytoskeleton (Cheek and Burgoyne, 1986). Two opposite roles have been described for F-actin: promoting vesicle transport (Gimenez-Molina et al, 2018;Lang et al, 2000;Trifaró et al, 2008) and preventing organelles, including vesicles, from reaching the PM (Toonen et al, 2006;Vitale et al, 1995Vitale et al, , 1991. Actin filaments are dynamic and showed a stimulation-dependent polymerization that has been suggested to facilitate regulated exocytosis (Trifaró et al, 1992;Vitale et al, 1991).…”

Spatial redistribution of neurosecretory vesicles upon stimulation accelerates their directed transport to the plasma membrane

“…Neuroendocrine chromaffin cells in the adrenal medulla are a model system to study the exocytosis of secretory vesicles (SVs) which is preceded by their transport from the Golgi apparatus to their site of exocytosis on the plasma membrane [ 1 ]. Defects in this process have been tied to a range of neurodegenerative diseases [ 2 ] by mechanisms that are not yet fully elucidated [ 3 ].…”

Spatial redistribution of neurosecretory vesicles upon stimulation accelerates their directed transport to the plasma membrane

“…Neuroendocrine chromaffin cells in the adrenal medulla are a model system to study the exocytosis of secretory vesicles (SVs) which is preceded by their transport from the Golgi apparatus to their site of exocytosis on the plasma membrane [1]. Defects in this process have been tied to a range of neurodegenerative diseases [2] by mechanism that…”

“…are not yet fully elucidated [3]. The highly crowded and stochastic nature of the cytoplasm, however, represents a major difficulty in exploring intracellular transport systems and as a result many of the mechanisms involved in bringing vesicles towards the plasma membrane remain elusive [1].…”

“…The copyright holder for this preprint (which this version posted May 20, 2021. ; https://doi.org/10.1101/2021.05.20.444910 doi: bioRxiv preprint mostly aligned radially while close to the cortex their density is lower and they align tangentially with the cortex [6]. In the periphery of the cell, transport along the actin filament network occurs through processive molecular motors such as myosin-V [8,9] [1,10] and non-processive such as Myosin-II [11]. Recruitment of SVs to the cortical actin network occurs via the action of Myosin-VI short-insert isoform which has both a processive and a non-processive function [12].…”