Protein Kinase A Activity Is Necessary for Fission and Fusion of Golgi to Endoplasmic Reticulum Retrograde Tubules

Tenorio, María J.; Luchsinger, Charlotte; Mardones, Gonzalo A.

doi:10.1371/journal.pone.0135260

Cited by 11 publications

(14 citation statements)

References 58 publications

(76 reference statements)

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“…Cells left untreated or treated with 5 μg/ml Brefeldin A (BFA) for 60 min at 37°C were analyzed by immunofluorescence microscopy as described previously [ 47 ]. Fixation of cells with methanol or 4% paraformaldehyde was performed when primary antibody reactivity demanded.…”

Section: Methodsmentioning

confidence: 99%

Distinct Biochemical Pools of Golgi Phosphoprotein 3 in the Human Breast Cancer Cell Lines MCF7 and MDA-MB-231

et al. 2016

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Golgi phosphoprotein 3 (GOLPH3) has been implicated in the development of carcinomas in many human tissues, and is currently considered a bona fide oncoprotein. Importantly, several tumor types show overexpression of GOLPH3, which is associated with tumor progress and poor prognosis. However, the underlying molecular mechanisms that connect GOLPH3 function with tumorigenicity are poorly understood. Experimental evidence shows that depletion of GOLPH3 abolishes transformation and proliferation of tumor cells in GOLPH3-overexpressing cell lines. Conversely, GOLPH3 overexpression drives transformation of primary cell lines and enhances mouse xenograft tumor growth in vivo. This evidence suggests that overexpression of GOLPH3 could result in distinct features of GOLPH3 in tumor cells compared to that of non-tumorigenic cells. GOLPH3 is a peripheral membrane protein mostly localized at the trans-Golgi network, and its association with Golgi membranes depends on binding to phosphatidylinositol-4-phosphate. GOLPH3 is also contained in a large cytosolic pool that rapidly exchanges with Golgi-associated pools. GOLPH3 has also been observed associated with vesicles and tubules arising from the Golgi, as well as other cellular compartments, and hence it has been implicated in several membrane trafficking events. Whether these and other features are typical to all different types of cells is unknown. Moreover, it remains undetermined how GOLPH3 acts as an oncoprotein at the Golgi. Therefore, to better understand the roles of GOLPH3 in cancer cells, we sought to compare some of its biochemical and cellular properties in the human breast cancer cell lines MCF7 and MDA-MB-231 with that of the non-tumorigenic breast human cell line MCF 10A. We found unexpected differences that support the notion that in different cancer cells, overexpression of GOLPH3 functions in diverse fashions, which may influence specific tumorigenic phenotypes.

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Section: Methodsmentioning

confidence: 99%

Distinct Biochemical Pools of Golgi Phosphoprotein 3 in the Human Breast Cancer Cell Lines MCF7 and MDA-MB-231

et al. 2016

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“…In particular, it has been shown that Golgi-to-ER retrograde transport plays a crucial role in the maintenance of Golgi morphology. Indeed, inhibition of this specific trafficking pathway results in a significant swelling of this organelle [32,[45][46][47][48][49]. To investigate whether the swelling of the Golgi by the inhibition of PSMD14 with CZM could be the result of the inhibition of the Golgi-to-ER retrograde transport, we first validated our findings in HeLa cells.…”

Section: Acute Inhibition Of the Deubiquitinating Enzyme Psmd14 Pertumentioning

confidence: 56%

“…Two other proteins that participates in this retrograde trafficking pathway, such as PKA and UVRAG [32,85], have been shown to be regulated by ubiquitination. Blockers of PKA signaling cause the inhibition of this trafficking pathway, a process that is accompanied by the swelling of the Golgi apparatus [32,49]. It has been shown that the catalytic PKA subunit (PKAc) is ubiquitinated by the CHIP E3 ligase, resulting in proteasomal degradation of PKAc and signaling shutdown [86].…”

Section: Discussionmentioning

confidence: 99%

The Proteasomal Deubiquitinating Enzyme PSMD14 Regulates Macroautophagy by Controlling Golgi-to-ER Retrograde Transport

Bustamante

Cereceda

González

et al. 2020

Cells

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Ubiquitination regulates several biological processes, however the role of specific members of the ubiquitinome on intracellular membrane trafficking is not yet fully understood. Here, we search for ubiquitin-related genes implicated in protein membrane trafficking performing a High-Content siRNA Screening including 1187 genes of the human “ubiquitinome” using amyloid precursor protein (APP) as a reporter. We identified the deubiquitinating enzyme PSMD14, a subunit of the 19S regulatory particle of the proteasome, specific for K63-Ub chains in cells, as a novel regulator of Golgi-to-endoplasmic reticulum (ER) retrograde transport. Silencing or pharmacological inhibition of PSMD14 with Capzimin (CZM) caused a robust increase in APP levels at the Golgi apparatus and the swelling of this organelle. We showed that this phenotype is the result of rapid inhibition of Golgi-to-ER retrograde transport, a pathway implicated in the early steps of the autophagosomal formation. Indeed, we observed that inhibition of PSMD14 with CZM acts as a potent blocker of macroautophagy by a mechanism related to the retention of Atg9A and Rab1A at the Golgi apparatus. As pharmacological inhibition of the proteolytic core of the 20S proteasome did not recapitulate these effects, we concluded that PSMD14, and the K63-Ub chains, act as a crucial regulatory factor for macroautophagy by controlling Golgi-to-ER retrograde transport.

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“…In particular, it has been shown that Golgi-to-ER retrograde transport plays a crucial role in the maintenance of Golgi morphology. Indeed, inhibition of this specific trafficking pathway results in a significant swelling of this organelle 40–45 . To investigate whether the swelling of the Golgi by the inhibition of PSMD14 with CZM could be the result of the inhibition of the Golgi-to-ER retrograde transport, we first validated our findings in HeLa cells.…”

Section: Resultsmentioning

confidence: 99%

“…Two other proteins that participates in this retrograde trafficking pathway, such as PKA and UVRAG 44, 76 , have been shown to be regulated by ubiquitination. Blockers of PKA signaling cause inhibition of this trafficking pathway, a process that is accompanied by the swelling of the Golgi apparatus 44, 45 . It has been shown that the catalytic PKA subunit (PKAc) is ubiquitinated by the CHIP E3 ligase, resulting in proteasomal degradation of PKAc and signaling shutdown 77 .…”

Section: Discussionmentioning

confidence: 99%

The proteasomal deubiquitinating enzyme PSMD14 regulates macroautophagy by controlling Golgi-to-ER retrograde transport

Bustamante

Cereceda

González

et al. 2020

Preprint

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31Ubiquitination regulates several biological processes. Here, we search for ubiquitin-related 32 genes implicated in protein membrane trafficking performing a High-Content siRNA 33 Screening including 1,187 genes of the human "ubiquitinome" using Amyloid Precursor 34 Protein (APP) as a reporter. We identified the deubiquitinating enzyme PSMD14, a subunit 35 of the 19S regulatory particle of the proteasome, specific for K63-Ub chains in cells, as a 36novel key regulator of Golgi-to-endoplasmic reticulum (ER) retrograde transport. Silencing 37 or pharmacological inhibition of PSMD14 caused a robust and rapid inhibition of Golgi-to-38 ER retrograde transport which leads to a potent blockage of macroautophagy by a 39 mechanism associated with the retention of Atg9A and Rab1A at the Golgi apparatus. 40Because pharmacological inhibition of the proteolytic core of the 20S proteasome did not 41 recapitulate these effects, we concluded that PSMD14, and their K-63-Ub chains, act as a 42 crucial regulator factor for macroautophagy by controlling Golgi-to-ER retrograde 43 transport. 44

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Protein Kinase A Activity Is Necessary for Fission and Fusion of Golgi to Endoplasmic Reticulum Retrograde Tubules

Cited by 11 publications

References 58 publications

Distinct Biochemical Pools of Golgi Phosphoprotein 3 in the Human Breast Cancer Cell Lines MCF7 and MDA-MB-231

Distinct Biochemical Pools of Golgi Phosphoprotein 3 in the Human Breast Cancer Cell Lines MCF7 and MDA-MB-231

The Proteasomal Deubiquitinating Enzyme PSMD14 Regulates Macroautophagy by Controlling Golgi-to-ER Retrograde Transport

The proteasomal deubiquitinating enzyme PSMD14 regulates macroautophagy by controlling Golgi-to-ER retrograde transport

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