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Martha S Cyert - One of the best experts on this subject based on the ideXlab platform.
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hph1 and hph2 are novel components of the sec63 sec62 posttranslational translocation complex that aid in vacuolar Proton ATPase biogenesis
Eukaryotic Cell, 2011Co-Authors: Francisco Pina, Allyson F Odonnell, Silvere Pagant, Hai Lan Piao, John P Miller, Stanley Fields, Elizabeth A Miller, Martha S CyertAbstract:Hph1 and Hph2 are homologous integral endoplasmic reticulum (ER) membrane proteins required for Saccharomyces cerevisiae survival under environmental stress conditions. To investigate the molecular functions of Hph1 and Hph2, we carried out a split-ubiquitin-membrane-based yeast two-hybrid screen and identified their interactions with Sec71, a subunit of the Sec63/Sec62 complex, which mediates posttranslational translocation of proteins into the ER. Hph1 and Hph2 likely function in posttranslational translocation, as they interact with other Sec63/Sec62 complex subunits, i.e., Sec72, Sec62, and Sec63. hph1Δ hph2Δ cells display reduced vacuole acidification; increased instability of Vph1, a subunit of vacuolar Proton ATPase (V-ATPase); and growth defects similar to those of mutants lacking V-ATPase activity. sec71Δ cells exhibit similar phenotypes, indicating that Hph1/Hph2 and the Sec63/Sec62 complex function during V-ATPase biogenesis. Hph1/Hph2 and the Sec63/Sec62 complex may act together in this process, as vacuolar acidification and Vph1 stability are compromised to the same extent in hph1Δ hph2Δ and hph1Δ hph2Δ sec71Δ cells. In contrast, loss of Pkr1, an ER protein that promotes posttranslocation assembly of Vph1 with V-ATPase subunits, further exacerbates hph1Δ hph2Δ phenotypes, suggesting that Hph1 and Hph2 function independently of Pkr1-mediated V-ATPase assembly. We propose that Hph1 and Hph2 aid Sec63/Sec62-mediated translocation of specific proteins, including factors that promote efficient biogenesis of V-ATPase, to support yeast cell survival during environmental stress.
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Hph1 and Hph2 Are Novel Components of the Sec63/Sec62 Posttranslational Translocation Complex That Aid in Vacuolar Proton ATPase Biogenesis
Eukaryotic Cell, 2010Co-Authors: Francisco Pina, Silvere Pagant, Hai Lan Piao, John P Miller, Stanley Fields, Elizabeth A Miller, Allyson F. O'donnell, Martha S CyertAbstract:Hph1 and Hph2 are homologous integral endoplasmic reticulum (ER) membrane proteins required for Saccharomyces cerevisiae survival under environmental stress conditions. To investigate the molecular functions of Hph1 and Hph2, we carried out a split-ubiquitin-membrane-based yeast two-hybrid screen and identified their interactions with Sec71, a subunit of the Sec63/Sec62 complex, which mediates posttranslational translocation of proteins into the ER. Hph1 and Hph2 likely function in posttranslational translocation, as they interact with other Sec63/Sec62 complex subunits, i.e., Sec72, Sec62, and Sec63. hph1Δ hph2Δ cells display reduced vacuole acidification; increased instability of Vph1, a subunit of vacuolar Proton ATPase (V-ATPase); and growth defects similar to those of mutants lacking V-ATPase activity. sec71Δ cells exhibit similar phenotypes, indicating that Hph1/Hph2 and the Sec63/Sec62 complex function during V-ATPase biogenesis. Hph1/Hph2 and the Sec63/Sec62 complex may act together in this process, as vacuolar acidification and Vph1 stability are compromised to the same extent in hph1Δ hph2Δ and hph1Δ hph2Δ sec71Δ cells. In contrast, loss of Pkr1, an ER protein that promotes posttranslocation assembly of Vph1 with V-ATPase subunits, further exacerbates hph1Δ hph2Δ phenotypes, suggesting that Hph1 and Hph2 function independently of Pkr1-mediated V-ATPase assembly. We propose that Hph1 and Hph2 aid Sec63/Sec62-mediated translocation of specific proteins, including factors that promote efficient biogenesis of V-ATPase, to support yeast cell survival during environmental stress.
Luis Carrasco - One of the best experts on this subject based on the ideXlab platform.
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The entry of reovirus into L cells is dependent on vacuolar Proton-ATPase activity.
Journal of Virology, 1996Co-Authors: C G Martínez, R Guinea, J Benavente, Luis CarrascoAbstract:Inhibitors of vacuolar Proton-ATPase activity (5 microM bafilomycin A1 or 50 nM concanamycin A) prevented infection by reovirus particles but not by infectious subviral particles (ISVPs). Neither compound affected virus attachment or internalization. However, both compounds potently blocked cleavage of the viral protein mu 1C. Finally, both reovirus particles and ISVPs efficiently translocated the toxin alpha-sarcin to the cytosol during virus entry. Bafilomycin A1 blocked translocation of alpha-sarcin by reovirus particles but not by ISVPs.
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requirement for vacuolar Proton ATPase activity during entry of influenza virus into cells
Journal of Virology, 1995Co-Authors: Rosario Guinea, Luis CarrascoAbstract:The role that endosomal acidification plays during influenza virus entry into MDCK cells has been analyzed by using the macrolide antibiotics bafilomycin A1 and concanamycin A as selective inhibitors of vacuolar Proton-ATPase (v-[H+]ATPase), the enzyme responsible for the acidification of endosomes. Bafilomycin A1 and concanamycin A, present at the low concentrations of 5 x 10(-7) and 5 x 10(-9) M, respectively, prevented the entry of influenza virus into cells when added during the first minutes of infection. Attachment of virion particles to the cell surface was not the target for the action of bafilomycin A1. N,N'-Dicyclohexylcarbodiimide, a nonspecific inhibitor of Proton-ATPases, also blocked virus entry, whereas elaiophylin, an inhibitor of the plasma-Proton ATPase, had no effect. The inhibitory actions of bafilomycin A1 and concanamycin A were tested in culture medium at different pHs. Both antibiotics powerfully prevented influenza virus infection when the virus was added under low-pH conditions. This inhibition was reduced if the virus was bound to cells at 4 degrees C prior to the addition of warm low-pH medium. Moreover, incubation of cells at acidic pH potently blocked influenza virus infection, even in the absence of antibiotics. These results indicate that a pH gradient, rather than low pH, is necessary for efficient entry of influenza virus into cells.
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Concanamycin A blocks influenza virus entry into cells under acidic conditions
FEBS Letters, 1994Co-Authors: Rosario Guinea, Luis CarrascoAbstract:Abstract The selective inhibitor of the vacuolar Proton-ATPase, concanamycin A, powerfully blocks influenza virus entry into cells, if present during the initial times of virus infection. Attachment of virus particles to cells is not prevented by concanamycin A, rather the exit of influenza virus from endosomes is the step blocked by this macrolide antibiotic. Inhibition of influenza virus entry into cells by concanamycin A or by nigericin takes place under acidic conditions. Moreover, if the pH gradient is abolished by pre-incubation of cells in acidic pH, influenza virus entry does not occur even in the absence of any inhibitors. These results indicate that acidic conditions per se are not sufficient to promote virus entry into cells; rather this step of virus infection requires a pH gradient.
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Concanamycin A: a powerful inhibitor of enveloped animal-virus entry into cells.
Biochemical and Biophysical Research Communications, 1994Co-Authors: Rosario Guinea, Luis CarrascoAbstract:Abstract Concanamycin A, a selective inhibitor of the vacuolar Proton ATPase, blocks the infection of animal cells by vesicular stomatitis virus, Semliki Forest virus and influenza virus even when the drug is present at the low concentration of 5 nM. Nevertheless the antibiotic prevents neither the attachment, to cells, of Semliki Forest virus nor its subsequent internalization. Under certain conditions, described in this communication, virus entry is prevented even when the pH of the medium is low, thus suggesting that a pH gradient, rather than low pH per se , is required to drive the entry, into cells, of these enveloped animal viruses.
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Entry of poliovirus into cells does not require a low-pH step.
Journal of Virology, 1993Co-Authors: Luis Perez, Luis CarrascoAbstract:Abstract The requirement of a low-pH step during poliovirus entry was investigated by using the macrolide antibiotic bafilomycin A1, which is a powerful and selective inhibitor of the vacuolar Proton-ATPases. Thus, viruses such as Semliki Forest virus and vesicular stomatitis virus that enter cells through endosomes and need their acidification, are potently inhibited by bafilomycin A1, whereas poliovirus infection is not affected by the antibiotic. The presence of lysosomotropic agents such as chloroquine, amantadine, dansylcadaverine, and monensin during poliovirus entry did not inhibit infection, further supporting the idea that poliovirus does not depend on a low-pH step to enter the cytoplasm. The effect of bafilomycin A1 on other members of the Picornaviridae family was also assayed. Encephalomyocarditis virus entry into HeLa cells was not affected by the macrolide antibiotic, whereas rhinovirus was sensitive. Coentry of toxins, such as alpha-sarcin, with viral particles was potently inhibited by bafilomycin A1, indicating that an active vacuolar Proton-ATPase is necessary for the early membrane permeabilization (coentry of alpha-sarcin) induced by poliovirus to take place.
Yoh Wada - One of the best experts on this subject based on the ideXlab platform.
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role of vacuolar type Proton ATPase in signal transduction
Biochimica et Biophysica Acta, 2015Co-Authors: Gehong Sunwada, Yoh WadaAbstract:Abstract The vacuolar H + -ATPase (V-ATPase) was first identified as an electrogenic Proton pump that acidifies the lumen of intra- and extracellular compartments. The acidic pH generated by V-ATPase is important for a wide range of cellular processes as well as acidification-independent processes such as secretion and membrane fusion. In addition to these housekeeping functions, recent studies implicate V-ATPase in the direct regulation and function of signaling pathways. In this review, we describe recent findings on the functions of V-ATPase in growth regulation and tissue physiology.
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Differential expression of a subunit isoforms of the vacuolar-type Proton pump ATPase in mouse endocrine tissues
Cell and Tissue Research, 2007Co-Authors: Ge-hong Sun-wada, Masamitsu Futai, Hiroyuki Tabata, Nobuyuki Kawamura, Yoh WadaAbstract:Vacuolar-type Proton ATPase (V-ATPase) is a multi-subunit enzyme that couples ATP hydrolysis to the translocation of Protons across membranes. Mammalian cells express four isoforms of the a subunit of V-ATPase. Previously, we have shown that V-ATPase with the a 3 isoform is highly expressed in pancreatic islets and is located in the membranes of insulin-containing granules in the β cells. The a 3 isoform functions in the regulation of hormone secretion. In this study, we have examined the distribution of a subunit isoforms in endocrine tissues, including the adrenal, parathyroid, thyroid, and pituitary glands, with isoform-specific antibodies. We have found that the a 3 isoform is strongly expressed in all these endocrine tissues. Our results suggest that functions of the a 3 isoform are commonly involved in the process of exocytosis in regulated secretion.
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lysosome and lysosome related organelles responsible for specialized functions in higher organisms with special emphasis on vacuolar type Proton ATPase
Cell Structure and Function, 2003Co-Authors: Gehong Sunwada, Yoh Wada, Masamitsu FutaiAbstract:Mammals contain various cells differentiated in both morphology and function, which play vital roles in tissue-specific functions. Late endosome/lysosome and lysosomal-related organelles are involved in these specialized functions including antigen presentation, bone remodeling and hormone regulation. To fulfill these diverse roles, lysosomes are present at different levels in different tissues and cell types; however, their morphology within these different tissues varies and the regulation of their activities differs with lysosomal compartments in some cells also functioning as secretory compartments. The luminal acidification of these organelles is closely correlated with their functions. This review will discuss the functions of lysosomes and lysosomal-related organelles, with particular emphasis on the major Proton pump, the vacuolar-type Proton ATPase (V-ATPase), which is responsible for luminal acidification.
Kouji Takeda - One of the best experts on this subject based on the ideXlab platform.
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effective mechanism for synthesis of neurotransmitter glutamate and its loading into synaptic vesicles
Neurochemical Research, 2017Co-Authors: Kouji Takeda, Tetsufumi UedaAbstract:Glutamate accumulation into synaptic vesicles is a pivotal step in glutamate transmission. This process is achieved by a vesicular glutamate transporter (VGLUT) coupled to v-type Proton ATPase. Normal synaptic transmission, in particular during intensive neuronal firing, would demand rapid transmitter re-filling of emptied synaptic vesicles. We have previously shown that isolated synaptic vesicles are capable of synthesizing glutamate from α-ketoglutarate (not from glutamine) by vesicle-bound aspartate aminotransferase for immediate uptake, in addition to ATP required for uptake by vesicle-bound glycolytic enzymes. This suggests that local synthesis of these substances, essential for glutamate transmission, could occur at the synaptic vesicle. Here we provide evidence that synaptosomes (pinched-off nerve terminals) also accumulate α-ketoglutarate-derived glutamate into synaptic vesicles within, at the expense of ATP generated through glycolysis. Glutamine-derived glutamate is also accumulated into synaptic vesicles in synaptosomes. The underlying mechanism is discussed. It is suggested that local synthesis of both glutamate and ATP at the presynaptic synaptic vesicle would represent an efficient mechanism for swift glutamate loading into synaptic vesicles, supporting maintenance of normal synaptic transmission.
Francisco Pina - One of the best experts on this subject based on the ideXlab platform.
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hph1 and hph2 are novel components of the sec63 sec62 posttranslational translocation complex that aid in vacuolar Proton ATPase biogenesis
Eukaryotic Cell, 2011Co-Authors: Francisco Pina, Allyson F Odonnell, Silvere Pagant, Hai Lan Piao, John P Miller, Stanley Fields, Elizabeth A Miller, Martha S CyertAbstract:Hph1 and Hph2 are homologous integral endoplasmic reticulum (ER) membrane proteins required for Saccharomyces cerevisiae survival under environmental stress conditions. To investigate the molecular functions of Hph1 and Hph2, we carried out a split-ubiquitin-membrane-based yeast two-hybrid screen and identified their interactions with Sec71, a subunit of the Sec63/Sec62 complex, which mediates posttranslational translocation of proteins into the ER. Hph1 and Hph2 likely function in posttranslational translocation, as they interact with other Sec63/Sec62 complex subunits, i.e., Sec72, Sec62, and Sec63. hph1Δ hph2Δ cells display reduced vacuole acidification; increased instability of Vph1, a subunit of vacuolar Proton ATPase (V-ATPase); and growth defects similar to those of mutants lacking V-ATPase activity. sec71Δ cells exhibit similar phenotypes, indicating that Hph1/Hph2 and the Sec63/Sec62 complex function during V-ATPase biogenesis. Hph1/Hph2 and the Sec63/Sec62 complex may act together in this process, as vacuolar acidification and Vph1 stability are compromised to the same extent in hph1Δ hph2Δ and hph1Δ hph2Δ sec71Δ cells. In contrast, loss of Pkr1, an ER protein that promotes posttranslocation assembly of Vph1 with V-ATPase subunits, further exacerbates hph1Δ hph2Δ phenotypes, suggesting that Hph1 and Hph2 function independently of Pkr1-mediated V-ATPase assembly. We propose that Hph1 and Hph2 aid Sec63/Sec62-mediated translocation of specific proteins, including factors that promote efficient biogenesis of V-ATPase, to support yeast cell survival during environmental stress.
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Hph1 and Hph2 Are Novel Components of the Sec63/Sec62 Posttranslational Translocation Complex That Aid in Vacuolar Proton ATPase Biogenesis
Eukaryotic Cell, 2010Co-Authors: Francisco Pina, Silvere Pagant, Hai Lan Piao, John P Miller, Stanley Fields, Elizabeth A Miller, Allyson F. O'donnell, Martha S CyertAbstract:Hph1 and Hph2 are homologous integral endoplasmic reticulum (ER) membrane proteins required for Saccharomyces cerevisiae survival under environmental stress conditions. To investigate the molecular functions of Hph1 and Hph2, we carried out a split-ubiquitin-membrane-based yeast two-hybrid screen and identified their interactions with Sec71, a subunit of the Sec63/Sec62 complex, which mediates posttranslational translocation of proteins into the ER. Hph1 and Hph2 likely function in posttranslational translocation, as they interact with other Sec63/Sec62 complex subunits, i.e., Sec72, Sec62, and Sec63. hph1Δ hph2Δ cells display reduced vacuole acidification; increased instability of Vph1, a subunit of vacuolar Proton ATPase (V-ATPase); and growth defects similar to those of mutants lacking V-ATPase activity. sec71Δ cells exhibit similar phenotypes, indicating that Hph1/Hph2 and the Sec63/Sec62 complex function during V-ATPase biogenesis. Hph1/Hph2 and the Sec63/Sec62 complex may act together in this process, as vacuolar acidification and Vph1 stability are compromised to the same extent in hph1Δ hph2Δ and hph1Δ hph2Δ sec71Δ cells. In contrast, loss of Pkr1, an ER protein that promotes posttranslocation assembly of Vph1 with V-ATPase subunits, further exacerbates hph1Δ hph2Δ phenotypes, suggesting that Hph1 and Hph2 function independently of Pkr1-mediated V-ATPase assembly. We propose that Hph1 and Hph2 aid Sec63/Sec62-mediated translocation of specific proteins, including factors that promote efficient biogenesis of V-ATPase, to support yeast cell survival during environmental stress.
