Genetically encoded fluorescent tools: Shining a little light on ER-to-Golgi transport

Seiler, Danette Kowal; Hay, Jesse

doi:10.1016/j.freeradbiomed.2022.03.004

Cited by 3 publications

(2 citation statements)

References 80 publications

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“…For the GFP-tagged VSV-G constructs, we employed (1) temperature-sensitive GFP-VSV-G ts045 which traffics following shift from 41 to 32 °C; (2) GFP-FM 4 -VSVG tm , which contains a luminal GFP and controlled aggregation domain connected to the WT VSV-G transmembrane domain and cytoplasmic C-terminus ( 8 ). Here, addition of the highly specific ligand D/D solubilizer causes dissociation of large cargo assemblies allowing packaging into COPII vesicles ( 28 , 29 ); (3) RUSH-VSV-G-GFP ( 30 , 31 ), which traffics after added biotin outcompetes binding of the construct to an ER-localized streptavidin “hook”. For the bulk-flow cargo, we employed GFP-FM 4 -hGH, an entirely luminal construct containing the controlled aggregation domain and hGH and no known functional sorting motif.…”

Section: Resultsmentioning

confidence: 99%

Steady-state regulation of COPII-dependent secretory cargo sorting by inositol trisphosphate receptors, calcium, and penta EF hand proteins

Held,

Lapka,

Sargeant

et al. 2023

Journal of Biological Chemistry

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Section: Resultsmentioning

confidence: 99%

Steady-state regulation of COPII-dependent secretory cargo sorting by inositol trisphosphate receptors, calcium, and penta EF hand proteins

Held,

Lapka,

Sargeant

et al. 2023

Journal of Biological Chemistry

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“…For the GFP-tagged VSV-G constructs, we employed: 1) temperature-sensitive GFP-VSV-G ts045 which traffics following shift from 41 to 32 °C; 2) GFP-FM 4 -VSVG tm , which contains a luminal GFP and controlled aggregation domain connected to the wildtype VSV-G transmembrane domain and cytoplasmic C-terminus (8). Here, addition of the highly specific ligand D/D Solubilizer causes dissociation of large cargo assemblies allowing packaging into COPII vesicles (28, 29). 3) RUSH-VSV-G-GFP (30, 31), which traffics after added biotin outcompetes binding of the construct to an ER-localized streptavidin “hook”.…”

Section: Resultsmentioning

confidence: 99%

Steady-State Regulation of COPII-Dependent Secretory Cargo Sorting by Inositol Trisphosphate Receptors, Calcium, and Penta EF Hand Proteins

Held

Hojanazarova

Sargeant

et al. 2020

Preprint

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ER Ca2+ regulates ER-to-Golgi transport machinery. Sustained Ca2+ signaling by inositol trisphosphate receptors (IP3Rs) leads to depression of cargo export through activation of penta EF hand protein (PEF) ALG-2 which reduces outer COPII coat at ER exit sites (ERES). However, we do not know whether tonic Ca2+ signals during steady-state conditions affect ER export rates and if so by what mechanisms. Here we report that partial depletion of IP3 receptors from NRK epithelial cells causes a marked increase of basal ER export of the transmembrane glycoprotein cargo VSV-G. The increased ER-to-Golgi transport required ALG-2 and was actuated by decreased peflin and increased ALG-2 at ER exit sites (ERES) – a condition previously demonstrated to stimulate COPII-dependent ER export. Upon IP3R depletion the amount of outer coat at ERES increased, the opposite to what occurs during ALG-2-dependent inhibition of secretion during agonist-driven Ca2+ signaling. The increased ER export correlated with reduced spontaneous cytosolic Ca2+ oscillations caused by the reduced number of Ca2+ release channels. IP3R depletion also unexpectedly resulted in partial depletion of ER luminal Ca2+ stores. The low Ca2+ conditions appeared to decrease both ALG-2 and peflin expression levels somewhat, but these were the only detectable expression changes in COPII trafficking machinery and the Ca2+ decrease had no detectable impact on ER stress. We conclude that at steady state, IP3Rs produce tonic Ca2+ signals that suppress the basal rate of ER export by maintaining lower outer coat targeting to ERES.

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The preamble to the free radical biology and medicine virtual special issue on “Targeting genetic biosensors to intracellular signaling pathways”

Eroğlu

Graier

Malli

2022

Free Radical Biology and Medicine

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Genetically encoded fluorescent tools: Shining a little light on ER-to-Golgi transport

Cited by 3 publications

References 80 publications

Steady-state regulation of COPII-dependent secretory cargo sorting by inositol trisphosphate receptors, calcium, and penta EF hand proteins

Steady-state regulation of COPII-dependent secretory cargo sorting by inositol trisphosphate receptors, calcium, and penta EF hand proteins

Steady-State Regulation of COPII-Dependent Secretory Cargo Sorting by Inositol Trisphosphate Receptors, Calcium, and Penta EF Hand Proteins

The preamble to the free radical biology and medicine virtual special issue on “Targeting genetic biosensors to intracellular signaling pathways”

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