Role of heterotrimeric GTP binding proteins in vesicular protein transport: indications for both classical and alternative G protein cycles

Helms, J. Bernd

doi:10.1016/0014-5793(95)00620-o

Cited by 84 publications

(60 citation statements)

References 81 publications

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“…It is not yet known whether G␣ subunits are active on intracellular membranes or whether the classical paradigm involving heptahelical receptors, G proteins, and effectors applies, because no specific receptors or effectors have been discovered to date on intracellular membranes (26).…”

Section: Discussionmentioning

confidence: 99%

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Identification and Characterization of GIV, a Novel Gαi/s -interacting Protein Found on COPI, Endoplasmic Reticulum-Golgi Transport Vesicles

Le-Niculescu

Niesman

Fischer

et al. 2005

Journal of Biological Chemistry

142

167

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In this report, we characterize GIV (G␣-interacting vesicle-associated protein), a novel protein that binds members of the G␣ i and G␣ s subfamilies of heterotrimeric G proteins. The G␣ interaction site was mapped to an 83-amino acid region of GIV that is enriched in highly charged amino acids. BLAST searches revealed two additional mammalian family members, Daple and an uncharacterized protein, FLJ00354. These family members share the highest homology at the G␣ binding domain, are homologous at the N terminus and central coiled coil domain but diverge at the C terminus. Using affinitypurified IgG made against two different regions of the protein, we localized GIV to COPI, endoplasmic reticulum (ER)-Golgi transport vesicles concentrated in the Golgi region in GH3 pituitary cells and COS7 cells. Identification as COPI vesicles was based on colocalization with ␤-COP, a marker for these vesicles. GIV also codistributes in the Golgi region with endogenous calnuc and the KDEL receptor, which are cis Golgi markers and with G␣ i3 -yellow fluorescent protein expressed in COS7 cells. By immunoelectron microscopy, GIV colocalizes with ␤-COP and G␣ i3 on vesicles found in close proximity to ER exit sites and to cis Golgi cisternae. In cell fractions prepared from rat liver, GIV is concentrated in a carrier vesicle fraction (CV2) enriched in ER-Golgi transport vesicles. ␤-COP and several G␣ subunits (G␣ i1-3 , G␣ s ) are also most enriched in CV2. Our results demonstrate the existence of a novel G␣-interacting protein associated with COPI transport vesicles that may play a role in G␣-mediated effects on vesicle trafficking within the Golgi and/or between the ER and the Golgi.

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Section: Discussionmentioning

confidence: 99%

“…Previously, heterotrimeric G proteins have been implicated in regulation of ER-Golgi transport (3,26), but the mechanisms involved are not yet understood. The discovery of a G␣-interacting protein located on these transport vesicles provides a new tool that may provide insights into the role of G␣ subunits in vesicular trafficking.…”

mentioning

confidence: 99%

Identification and Characterization of GIV, a Novel Gαi/s -interacting Protein Found on COPI, Endoplasmic Reticulum-Golgi Transport Vesicles

Le-Niculescu

Niesman

Fischer

et al. 2005

Journal of Biological Chemistry

142

167

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“…Besides their plasma membrane location, heterotrimeric G proteins also are found on membranes of intracellular compartments along both the endocytic and secretory pathways where indirect evidence suggests they play several roles in membrane trafficking (Bomsel and Mostov, 1992;Helms, 1995;Nurnberg and Ahnert-Hilger, 1996;Stow and Heimann, 1998). One of the prototypical heterotrimeric G proteins, G␣s, the stimulatory subunit of heterotrimeric G proteins, has been suggested to regulate endocytic trafficking.…”

Section: Introductionmentioning

confidence: 99%

Regulation of Epidermal Growth Factor Receptor Degradation by Heterotrimeric Gαs Protein

Zheng

Lavoie

Tang

et al. 2004

MBoC

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Heterotrimeric G proteins have been implicated in the regulation of membrane trafficking, but the mechanisms involved are not well understood. Here, we report that overexpression of the stimulatory G protein subunit (Gαs) promotes ligand-dependent degradation of epidermal growth factor (EGF) receptors and Texas Red EGF, and knock-down of Gαs expression by RNA interference (RNAi) delays receptor degradation. We also show that Gαs and its GTPase activating protein (GAP), RGS-PX1, interact with hepatocyte growth factor-regulated tyrosine kinase substrate (Hrs), a critical component of the endosomal sorting machinery. Gαs coimmunoprecipitates with Hrs and binds Hrs in pull-down assays. By immunofluorescence, exogenously expressed Gαs colocalizes with myc-Hrs and GFP-RGS-PX1 on early endosomes, and expression of either Hrs or RGS-PX1 increases the localization of Gαs on endosomes. Furthermore, knock-down of both Hrs and Gαs by double RNAi causes greater inhibition of EGF receptor degradation than knock-down of either protein alone, suggesting that Gαs and Hrs have cooperative effects on regulating EGF receptor degradation. These observations define a novel regulatory role for Gαs in EGF receptor degradation and provide mechanistic insights into the function of Gαs in endocytic sorting

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“…There is compelling evidence that caveolin-1 and caveolin-2 are present at the Golgi complex (Luetterforst et al, 1999;Mora et al, 1999;Parolini et al, 1999;Machleidt et al, 2000). In addition to caveolin, heterotrimeric G proteins have been localized to the Golgi complex, and several lines of evidence support a role in intracellular protein transport (Bomsel and Mostov, 1992;Helms, 1995). More recently, heterotrimeric G proteins have been implicated in the regulation of Golgi structure (Jamora et al, 1997;Yamaguchi et al, 1997).…”

Section: Introductionmentioning

confidence: 99%

Sphingomyelin-enriched Microdomains at the Golgi Complex

Gkantiragas

Brügger

Stüven

et al. 2001

MBoC

155

134

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Sphingomyelin- and cholesterol-enriched microdomains can be isolated as detergent-resistant membranes from total cell extracts (total-DRM). It is generally believed that this total-DRM represents microdomains of the plasma membrane. Here we describe the purification and detailed characterization of microdomains from Golgi membranes. These Golgi-derived detergent-insoluble complexes (GICs) have a low buoyant density and are highly enriched in lipids, containing 25% of total Golgi phospholipids including 67% of Golgi-derived sphingomyelin, and 43% of Golgi-derived cholesterol. In contrast to total-DRM, GICs contain only 10 major proteins, present in nearly stoichiometric amounts, including the α- and β-subunits of heterotrimeric G proteins, flotillin-1, caveolin, and subunits of the vacuolar ATPase. Morphological data show a brefeldin A-sensitive and temperature-sensitive localization to the Golgi complex. Strikingly, the stability of GICs does not depend on its membrane environment, because, after addition of brefeldin A to cells, GICs can be isolated from a fused Golgi-endoplasmic reticulum organelle. This indicates that GIC microdomains are not in a dynamic equilibrium with neighboring membrane proteins and lipids. After disruption of the microdomains by cholesterol extraction with cyclodextrin, a subcomplex of several GIC proteins including the B-subunit of the vacuolar ATPase, flotillin-1, caveolin, and p17 could still be isolated by immunoprecipitation. This indicates that several of the identified GIC proteins localize to the same microdomains and that the microdomain scaffold is not required for protein interactions between these GIC proteins but instead might modulate their affinity.

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Role of heterotrimeric GTP binding proteins in vesicular protein transport: indications for both classical and alternative G protein cycles

Cited by 84 publications

References 81 publications

Identification and Characterization of GIV, a Novel Gαi/s -interacting Protein Found on COPI, Endoplasmic Reticulum-Golgi Transport Vesicles

Identification and Characterization of GIV, a Novel Gαi/s -interacting Protein Found on COPI, Endoplasmic Reticulum-Golgi Transport Vesicles

Regulation of Epidermal Growth Factor Receptor Degradation by Heterotrimeric Gαs Protein

Sphingomyelin-enriched Microdomains at the Golgi Complex

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