Actin filaments are involved in the maintenance of Golgi cisternae morphology and intra‐Golgi pH

Abstract

Here we examine the contribution of actin dynamics to the architecture and pH of the Golgi complex. To this end, we have used toxins that depolymerize (cytochalasin D, latrunculin B, mycalolide B, and Clostridium botulinum C2 toxin) or stabilize (jasplakinolide) filamentous actin. When various clonal cell lines were examined by epifluorescence microscopy, all of these actin toxins induced compaction of the Golgi complex. However, ultrastructural analysis by transmission electron microscopy and electron tomography/three-dimensional modelling of the Golgi complex showed that F-actin depolymerization first induces perforation/fragmentation and severe swelling of Golgi cisternae, which leads to a completely disorganized structure. In contrast, F-actin stabilization results only in cisternae perforation/fragmentation. Concomitantly to actin depolymerization-induced cisternae swelling and disorganization, the intra-Golgi pH significantly increased. Similar ultrastructural and Golgi pH alkalinization were observed in cells treated with the vacuolar H þ -ATPases inhibitors bafilomycin A1 and concanamycin A. Overall, these results suggest that actin filaments are implicated in the preservation of the flattened shape of Golgi cisternae. This maintenance seems to be mediated by the regulation of the state of F-actin assembly on the Golgi pH homeostasis. Cell Motil. Cytoskeleton 63: 778-791, 2006. '

Actin Filaments Are Involved in the Coupling of V0-V1 Domains of Vacuolar H+-ATPase at the Golgi Complex

“…Actin-depolymerizing Agents Induce the V 1 -V 0 Disassociation of the V-ATPase at the Golgi Complex-On the one hand, we previously reported that actin depolymerization raises the pH in the Golgi and alters Golgi-to-ER and post-Golgi trafficking (47)(48)(49). These results are similar to those recorded after the pharmacological inhibition of V-ATPase using bafilomycin and concanamycin (44).…”

“…Knowing that B and C subunits of the V 1 domain bind to F-and/or G-actin (25,26,28,50), we hypothesized that actin could participate in Golgi pH homeostasis through the regulation of V-ATPase activity. In particular, we hypothesized that microfilaments are crucial to the maintenance of V 0 and V 1 domain association (49,51). In this study, we provide experimental evidence that microfilaments do indeed maintain V 0 and V 1 domain association via two mechanisms as follows: one through the interaction between actin and subunits B2 and C1, and other based on the actin-dependent integrity of detergent-resistant membranes (DRMs) where V-ATPase localizes.…”

“…In contrast, much less is known about the subcellular localization and regulatory mechanisms of V-ATPase in the Golgi. Our group previously reported significant similarities between events occurring after microfilament disruption (with actin-depolymerizing agents) (47)(48)(49) and those seen after the pharmacological inhibition of V-ATPase (with bafilomycin A1 and concanamycin A) (43,44,49). These similarities include the following: (a) membrane trafficking alterations in the Golgi-to-ER and post-Golgi protein transports; (b) alkalization of the Golgi complex, and (c) strong dilatation of cisternae, observed under the electron microscope.…”

Phosphatidylinositol- and phosphatidylcholine-transfer activity of PITPβ is essential for COPI-mediated retrograde transport from the Golgi to the endoplasmic reticulum

“…The Golgi morphology described here is reminiscent of that caused by a number of agents that impact on cytoskeletal architecture including latrunculin B, cytochalasin D and disruption of the cytoskeletal anchor, Syne-1 (Valderrama et al, 1998;Valderrama et al, 2001;Gough and Beck, 2004;Lazaro-Dieguez et al, 2006). Syne-1 localises to the Golgi (Gough et al, 2003) and expression of fragments from Syne-1 alters the structure of the Golgi complex, which collapses into a compact juxtanuclear structure (Gough and Beck, 2004).…”

Dynamics of an F-actin aggresome generated by the actin-stabilizing toxin jasplakinolide

“…Control and treated Vero cells were processed as previously described (Lazaro-Dieguez et al, 2006). Ultrathin sections were stained with 2% uranyl acetate for 30 minutes, then with lead citrate for 10 minutes and observed with a JEOL 1010 transmission electron microscope operating at 80 kV and provided with a Gatan BioScan model 792 module for acquisition of digital images with Digital Micrograph 3.4.3 acquisition software (Gatan, Pleasanton, CA).…”