Analysis of Arf GTP‐Binding Protein Function in Cells

Cohen, Lee Ann; Donaldson, Julie G.

doi:10.1002/0471143030.cb1412s48

Cited by 21 publications

(26 citation statements)

References 15 publications

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“…S7A), but Arf1 activity remained elevated, vesicle uncoating was impaired, trafficking was delayed and Golgi ribbons were dispersed. Activation of Gi and release of ‘free’ Gβγ dimers is also a central mechanism by which GIV terminates Arf1 at/near acceptor (i.e., Golgi) membranes, and is consistent with prior work demonstrating the inhibitory role of Gβγ in Arf signaling (Cohen and Donaldson, 2010; Colombo et al, 1995). Although hierarchically dominant, the GIV→Gβγ pathway is dependent on the GIV→ArfGAP2/3 pathway, because the ability of GIV-GEF to downregulate Arf1 activity is abolished in cells without ArfGAP3 (Fig.…”

Section: Discussionsupporting

confidence: 89%

“…To determine if GIV or its GEF function modulate Arf1 activity in cells, we used the previously validated pull-down assay with a GST-GAT domain of GGA3 which selectively binds the active GTP-bound pool of Arf1 (Cohen and Donaldson, 2010). We found that compared to control cells, the levels of Arf1•GTP were increased ~60% in GIV-depleted cells (Fig.…”

Section: Resultsmentioning

confidence: 99%

“…Purification of GST-GAT protein and assessment of Arf1 activation were described previously (Cohen and Donaldson, 2010). Immunofluorescence and confocal microscopy was performed as described previously (Ghosh et al, 2010).…”

Section: Methodsmentioning

confidence: 99%

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Activation of Gαi at the Golgi by GIV/Girdin Imposes Finiteness in Arf1 Signaling

et al. 2015

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SUMMARY A long-held tenet of heterotrimeric G protein signal transduction is that it is triggered by G-protein-coupled receptors (GPCRs) at the PM. Here we demonstrate that Gi is activated in the Golgi by GIV/Girdin, a non-receptor guanine-nucleotide exchange factor (GEF). GIV-dependent activation of Gi at the Golgi maintains the finiteness of the cyclical activation of ADP-ribosylation factor 1 (Arf1), a fundamental step in vesicle traffic in all eukaryotes. Several interactions with other major components of Golgi trafficking, e.g., active Arf1, its regulator, ArfGAP2/3, and the adaptor protein β-COP enable GIV to coordinately regulate Arf1 signaling. When the GIV-Gαi pathway is selectively inhibited, levels of GTP-bound Arf1 are elevated and protein transport along the secretory pathway is delayed. These findings define a paradigm in non-canonical G protein signaling at the Golgi which places GIV-GEF at the crossroads between signals gated by the trimeric G proteins and the Arf-family of monomeric GTPases.

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Section: Discussionsupporting

confidence: 89%

Section: Resultsmentioning

confidence: 99%

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Activation of Gαi at the Golgi by GIV/Girdin Imposes Finiteness in Arf1 Signaling

et al. 2015

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“…That GIV terminates Arf1 at/near acceptor (i.e. Golgi) membranes via release of free Gbg is consistent with prior work demonstrating the inhibitory role of Gbg in Arf signaling [78,79]. Thus, this work elucidated some of poorly understood functions of G proteins on the Golgi and illuminated the role of trimeric G protein signaling in the secretory pathway.…”

Section: Giv Regulates Secretion By Activating Gai On Golgi Membranessupporting

confidence: 86%

Heterotrimeric G protein signaling via GIV/Girdin: Breaking the rules of engagement, space, and time

et al. 2016

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Canonical signal transduction via heterotrimeric G proteins is spatially and temporally restricted, i.e., triggered exclusively at the plasma membrane (PM), only by agonist activation of G-protein-coupled receptors (GPCRs) via a process that completes within a few hundred milliseconds. Recently, a rapidly emerging paradigm has revealed a non-canonical pathway for activation of trimeric G proteins by the non-receptor guanidine-nucleotide exchange factor (GEF), GIV/Girdin. This pathway has distinctive temporal and spatial features and an unusual profile of receptor engagement: Diverse classes of receptors, not just GPCRs can engage with GIV-GEF to trigger such activation. Such activation is spatially and temporally unrestricted, i.e., can occur both at the PM and on internal membranes discontinuous with the PM, and can continue for prolonged periods of time. Here we review the molecular mechanisms that govern non-canonical G protein activation by GIV-GEF and the relevance of this new paradigm in health and disease.

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“…ARF activity was measured using an ARF pull-down assay essentially as described earlier 70 . GST-VHS-GAT (VHS-GAT from GGA3) was bacterially expressed, and the cell pellet spun down at 4000 rpm −1 .…”

Section: Methodsmentioning

confidence: 99%

A CREB3–ARF4 signalling pathway mediates the response to Golgi stress and susceptibility to pathogens

et al. 2013

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SUMMARY Treatment of cells with Brefeldin A (BFA) blocks secretory vesicle transport and causes a collapse of the Golgi apparatus. To gain more insight into the cellular mechanisms mediating BFA toxicity, we conducted a genome-wide haploid genetic screen that led to the identification of the small G protein ADP-ribosylation factor 4 (ARF4). ARF4 depletion preserves viability, Golgi integrity and cargo trafficking in the presence of BFA, and these effects depend on the guanine nucleotide exchange factor GBF1 and other ARF isoforms including ARF1 and ARF5. ARF4 knockdown cells show increased resistance to several human pathogens including Chlamydia trachomatis and Shigella flexneri. Furthermore, ARF4 expression is induced when cells are exposed to several Golgi-disturbing agents and requires the CREB3/Luman transcription factor whose downregulation mimics ARF4 loss. Thus, we have uncovered a CREB3–ARF4 signaling cascade that may be part of a Golgi stress response set in motion by stimuli compromising Golgi capacity.

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Analysis of Arf GTP‐Binding Protein Function in Cells

Cited by 21 publications

References 15 publications

Activation of Gαi at the Golgi by GIV/Girdin Imposes Finiteness in Arf1 Signaling

Activation of Gαi at the Golgi by GIV/Girdin Imposes Finiteness in Arf1 Signaling

Heterotrimeric G protein signaling via GIV/Girdin: Breaking the rules of engagement, space, and time

A CREB3–ARF4 signalling pathway mediates the response to Golgi stress and susceptibility to pathogens

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