Vacuolar ATPase inactivation blocks recycling to the <i>trans</i>‐Golgi network from the plasma membrane

Reaves, Barbara J.; Banting, George

doi:10.1016/0014-5793(94)00437-4

Cited by 71 publications

(60 citation statements)

References 29 publications

(19 reference statements)

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“…7 Bafilomycin A 1 does not inhibit the fusion between autophagosomes and endosomes. 17 The internal pH of acidic compartments may be important in the fusion of lysosomes. But it is not clear if the internal pH of lysosomes or of autophagosomes regulates the autophagosome-lysosome fusion, partly because the internal pH of autophagosomes is not known.…”

Section: Discussionmentioning

confidence: 99%

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Autophagosome-Lysosome Fusion Depends on the pH in Acidic Compartments in CHO Cells

Kawai¹,

Uchiyama²,

Takano³

et al. 2007

Autophagy

196

158

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AcKNOwledgemeNTSWe are indebted to Prof. Mori M. (Kumamoto University, Japan) for generously providing pCAGGS-pOTC-GFP. We thank Ms. Nagami Yamashita for her supports. Prof. Ohkuma died on November 5, 2006 by myocardial infarction at his home. We greatly miss him as a scientist and a friend. We offer sincere thanks to all the friends, colleagues and former collaborators of Prof. Ohkuma who showed him kindness during his lifetime. AbSTrAcTAutophagy is the bulk degradation of cytoplasmic constituents in response to starvation and other environmental or intracellular cues. During this process, most of the cytoplasm is sequestered into autophagosomes, which then fuse with lysosomes where the degradation of the sequestered material proceeds. We investigated the relationship between autophagosome-lysosome fusion and the pH in acidic compartments by visualizing the fusion process using fluorescence in CHO cells. In this experiment, mitochondria were labeled with GFP by transfecting CHO cells with the presequence of ornithine transcarbamylase, and lysosomes were labeled with Texas Red Dextran; any fusion was identified by the colocalization of mitochondria (in autophagosomes) and lysosomes using fluorescence microscopy. When CHO cells were treated with rapamycin or starvation medium to induce autophagy, the colocalization of fluorescence was observed. Whereas when they were treated with 3-MA, an inhibitor of autophagy, the colocalization disappeared. We conclude that the colocalization reflects the fusion of autophagosomes and lysosomes. Moreover, when the CHO cells were treated with drugs that increase the pH of acidic compartments, the colocalization disappeared. This suggests that the autophagosome-lysosome fusion is inhibited by increasing pH in acidic compartments independently of V-ATPase activity in CHO cells.

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Section: Discussionmentioning

confidence: 99%

“…[14][15][16][17][18] Furthermore, the activity of V-ATPase is required for the maturation of endosomes Figure 1. Autophagy and the colocalization of mitochondria and lysosomes.…”

Section: Discussionmentioning

confidence: 99%

Autophagosome-Lysosome Fusion Depends on the pH in Acidic Compartments in CHO Cells

Kawai¹,

Uchiyama²,

Takano³

et al. 2007

Autophagy

196

158

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“…The Golgi apparatus events were interfered with by using the macrolide antibiotic BAF, an inhibitor of vacuolar proton ATPase activity along the Golgi apparatus cisternae (15), and WO, a potent inhibitor of the phosphatidylinositol 3-kinase activity involved in trafficking at the trans-Golgi apparatus level (32). The pH gradient in the Golgi apparatus was also disrupted by treatment with CQ, an acidotropic compound (40). As shown in Fig.…”

Section: Vol 85 2011 Role Of Cholesterol In Hbv Assembly 13375mentioning

confidence: 99%

Cholesterol Depletion of Hepatoma Cells Impairs Hepatitis B Virus Envelopment by Altering the Topology of the Large Envelope Protein

Dorobantu

Macovei

Lazar

et al. 2011

J Virol

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Previous reports have shown that cholesterol depletion of the membrane envelope of the hepatitis B virus (HBV) impairs viral infection of target cells.A potential function of this lipid in later steps of the viral life cycle remained controversial, with secretion of virions and subviral particles (SVP) being either inhibited or not affected, depending on the experimental approach employed to decrease the intracellular cholesterol level. This work addressed the role of host cell cholesterol on HBV replication, assembly, and secretion, using an alternative method to inhibition of the enzymes involved in the biosynthesis pathway. Growing HBV-producing cells with lipoprotein-depleted serum (LPDS) resulted in an important reduction of the amount of cholesterol within 24 h of treatment (about 40%). Cell exposure to chlorpromazine, an inhibitor of the clathrin-mediated pathway used by the low-density lipoprotein receptor for endocytosis, also impacted the cholesterol level; however, this level of inhibition was not achievable when the synthesis inhibitor lovastatin was used. HBV secretion was significantly inhibited in cholesterol-depleted cells (by ϳ80%), while SVP release remained unaffected. The viral DNA genome accumulated in LPDS-treated cells in a time-dependent manner. Specific immunoprecipitation of nucleocapsids and mature virions revealed an increased amount of naked nucleocapsids, while synthesis of the envelope proteins occurred as normally. Following analysis of the large envelope protein conformation in purified microsomes, we concluded that cholesterol is important in maintaining the dual topology of this polypeptide, which is critical for viral envelopment.A large variety of viruses, of which many are important human pathogens, depend on lipid and cholesterol metabolism in host cells during at least one step of their life cycle. Hepatitis C virus, for instance, relies on lipids for entry into target cells (21), RNA replication (22), viral assembly (45), as well as infectivity (1, 37). Cholesterol-rich plasma membrane domains (lipid rafts) are important for HIV entry, assembly, and infectivity (2). In the case of hepatitis B virus (HBV), efficient infection of hepatocytes was shown to be dependent on the cholesterol content of the viral envelope (6, 37); more recently, a role for caveolin-1, a structural protein of lipid rafts, was suggested in HBV entry (30).HBV is an enveloped member of the Hepadnaviridae family bearing an unusual feature among animal viruses, in that multiple types of virus-related particles are assembled in infected cells. The infectious virions, also called "Dane" particles, are sphere-shaped, 42-nm-diameter particles containing the nucleocapsid surrounded by an envelope composed of cellular lipids and three structural viral proteins. These are designated the large (L), middle (M), and small (S) proteins and derive from the same open reading frame, sharing a common S domain (43). In addition to mature virions, coreless, noninfectious lipoprotein particles occurring in two morphologi...

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“…V-ATPase inhibition does, however, inhibit activation or dissociation of these agents from their receptors in a pH-dependent step after endocytosis has taken place. Deacidification of vesicles has also been demonstrated to inhibit the subsequent traffic of a number of specific proteins (Chapman and Munro, 1994;Clague et al, 1994;Reaves and Banting, 1994). Granule cells loaded with D-[5Hlaspartate for 60 mm were subjected to field stimulation at the indicated frequency (four trains of 400 pulses separated by 10 s between trains).…”

Section: Synaptic Vesicle Recycling Does Not Require Lumenal Acidificmentioning

confidence: 99%

Synaptic Vesicle Recycling in Cultured Cerebellar Granule Cells: Role of Vesicular Acidification and Refilling

Cousin

Nicholls

1997

Journal of Neurochemistry

106

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Abstract:The role of the transvesicular protonmotive force in synaptic vesicle recycling was investigated in cultured cerebellar granule cells. The vesicular V-ATPase was inhibited by 1 ,uM bafilomycin Al; as an alternative, the pH component of the gradient was selectively collapsed by equilibration of the cells with 10 mM methylamine and monitored with the fluorescent probe Lysosensor Green. Electrical field-evoked exocytosis of D-[ 3H]aspartate was inhibited by bafilomycin Al but not by methylamine, indicating that a transvesicular membrane potential rather than pH gradient is required for transmitter retention within vesicles. In contrast, neither compound affected the field-evoked uptake, recycling, or destaining of the vesicle-specific dye FM2-10; thus, vesicles whose lumens were neutral and/or depleted of transmitter could still recycle in the nerve terminal. No exhaustion of o-[3H]aspartate exocytosis was observed when cells were subjected to six consecutive trains of field stimuli (40 Hz/l0 s separated by 10 s). In contrast, the release of preloaded FM2-10 was reduced by -~50%, with each stimulus indicating that unlabeled vesicles with accumulated D-[3H]aspartate were competing with labeled vesides for exocytosis. As D-[3HI aspartate was accumulated rapidly across the vesicle membrane from the large cytoplasmic pool, the transmitter-loaded but unlabelled vesides may represent refilled recycling vesicles. FM2-l 0 destaining and D-[3H]aspartate exocytosis were reduced in parallel at low frequencies, challenging a role for transient vesicle fusion. Key Words: Exocytosis-Vesicle-Refilling-D-[3H]Aspartate-FM1 -43-Granule cell.

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Vacuolar ATPase inactivation blocks recycling to the trans‐Golgi network from the plasma membrane

Cited by 71 publications

References 29 publications

Autophagosome-Lysosome Fusion Depends on the pH in Acidic Compartments in CHO Cells

Autophagosome-Lysosome Fusion Depends on the pH in Acidic Compartments in CHO Cells

Cholesterol Depletion of Hepatoma Cells Impairs Hepatitis B Virus Envelopment by Altering the Topology of the Large Envelope Protein

Synaptic Vesicle Recycling in Cultured Cerebellar Granule Cells: Role of Vesicular Acidification and Refilling

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