Functional Expression and Characterization of Schizosaccharomyces pombe Avt3p as a Vacuolar Amino Acid Exporter in Saccharomyces cerevisiae
In Saccharomyces cerevisiae, Avt3p and Avt4p mediate the extrusion of several amino acids from the vacuolar lumen into the cytosol. SpAvt3p of Schizosaccharomyces pombe, a homologue of these vacuolar amino acid transporters, has been indicated to be involved in spore formation. In this study, we con...
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| creator | Chardwiriyapreecha, Soracom Manabe, Kunio Iwaki, Tomoko Kawano-Kawada, Miyuki Sekito, Takayuki Lunprom, Siriporn Akiyama, Koichi Takegawa, Kaoru Kakinuma, Yoshimi |
| description | In Saccharomyces cerevisiae, Avt3p and Avt4p mediate the extrusion of several amino acids from the vacuolar lumen into the cytosol. SpAvt3p of Schizosaccharomyces pombe, a homologue of these vacuolar amino acid transporters, has been indicated to be involved in spore formation. In this study, we confirmed that GFP-SpAvt3p localized to the vacuolar membrane in S. pombe. The amounts of various amino acids increased significantly in the vacuolar pool of avt3Δ cells, but decreased in that of avt3+-overexpressing avt3Δ cells. These results suggest that SpAvt3p participates in the vacuolar compartmentalization of amino acids in S. pombe. To examine the export activity of SpAvt3p, we expressed the avt3+ gene in S. cerevisiae cells. We found that the heterologously overproduced GFP-SpAvt3p localized to the vacuolar membrane in S. cerevisiae. Using the vacuolar membrane vesicles isolated from avt3+-overexpressing S. cerevisiae cells, we detected the export activities of alanine and tyrosine in an ATP-dependent manner. These activities were inhibited by the addition of a V-ATPase inhibitor, concanamycin A, thereby suggesting that the activity of SpAvt3p is dependent on a proton electrochemical gradient generated by the action of V-ATPase. In addition, the amounts of various amino acids in the vacuolar pools of S. cerevisiae cells were decreased by the overproduction of SpAvt3p, which indicated that SpAvt3p was functional in S. cerevisiae cells. Thus, SpAvt3p is a vacuolar transporter that is involved in the export of amino acids from S. pombe vacuoles. |
| doi_str_mv | 10.1371/journal.pone.0130542 |
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SpAvt3p of Schizosaccharomyces pombe, a homologue of these vacuolar amino acid transporters, has been indicated to be involved in spore formation. In this study, we confirmed that GFP-SpAvt3p localized to the vacuolar membrane in S. pombe. The amounts of various amino acids increased significantly in the vacuolar pool of avt3Δ cells, but decreased in that of avt3+-overexpressing avt3Δ cells. These results suggest that SpAvt3p participates in the vacuolar compartmentalization of amino acids in S. pombe. To examine the export activity of SpAvt3p, we expressed the avt3+ gene in S. cerevisiae cells. We found that the heterologously overproduced GFP-SpAvt3p localized to the vacuolar membrane in S. cerevisiae. Using the vacuolar membrane vesicles isolated from avt3+-overexpressing S. cerevisiae cells, we detected the export activities of alanine and tyrosine in an ATP-dependent manner. These activities were inhibited by the addition of a V-ATPase inhibitor, concanamycin A, thereby suggesting that the activity of SpAvt3p is dependent on a proton electrochemical gradient generated by the action of V-ATPase. In addition, the amounts of various amino acids in the vacuolar pools of S. cerevisiae cells were decreased by the overproduction of SpAvt3p, which indicated that SpAvt3p was functional in S. cerevisiae cells. Thus, SpAvt3p is a vacuolar transporter that is involved in the export of amino acids from S. pombe vacuoles.</description><identifier>ISSN: 1932-6203</identifier><identifier>EISSN: 1932-6203</identifier><identifier>DOI: 10.1371/journal.pone.0130542</identifier><identifier>PMID: 26083598</identifier><language>eng</language><publisher>United States: Public Library of Science</publisher><subject>Adenosine triphosphatase ; Adenosine Triphosphatases - metabolism ; Alanine ; Amino Acid Sequence ; Amino Acid Transport Systems - genetics ; Amino Acid Transport Systems - metabolism ; Amino acids ; Amino Acids - metabolism ; ATPases ; Biological Transport ; Concanamycin A ; Corrosion inhibitors ; Cytosol ; Electrochemistry ; Eukaryotes ; Exports ; Extrusion ; Genes ; Genetics ; H+-transporting ATPase ; Homology ; Immunoblotting ; Intracellular Membranes ; Laboratories ; Membrane vesicles ; Molecular Sequence Data ; Physiology ; Proteins ; Saccharomyces cerevisiae ; Saccharomyces cerevisiae - genetics ; Saccharomyces cerevisiae - growth & development ; Saccharomyces cerevisiae - metabolism ; Schizosaccharomyces - metabolism ; Schizosaccharomyces pombe ; Schizosaccharomyces pombe Proteins - genetics ; Schizosaccharomyces pombe Proteins - metabolism ; Sequence Homology, Amino Acid ; Tyrosine ; Vacuoles ; Vacuoles - metabolism ; Yeast</subject><ispartof>PloS one, 2015-06, Vol.10 (6), p.e0130542-e0130542</ispartof><rights>COPYRIGHT 2015 Public Library of Science</rights><rights>2015 Chardwiriyapreecha et al. This is an open access article distributed under the terms of the Creative Commons Attribution License: http://creativecommons.org/licenses/by/4.0/ (the “License”), which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited Notwithstanding the ProQuest Terms and Conditions, you may use this content in accordance with the terms of the License.</rights><rights>2015 Chardwiriyapreecha et al 2015 Chardwiriyapreecha et al</rights><lds50>peer_reviewed</lds50><oa>free_for_read</oa><woscitedreferencessubscribed>false</woscitedreferencessubscribed><citedby>FETCH-LOGICAL-c758t-8fb27fea8af9cfa9f681b8f64b715d24f4b52e9efdc4ed035bdede3f585001833</citedby><cites>FETCH-LOGICAL-c758t-8fb27fea8af9cfa9f681b8f64b715d24f4b52e9efdc4ed035bdede3f585001833</cites></display><links><openurl>$$Topenurl_article</openurl><openurlfulltext>$$Topenurlfull_article</openurlfulltext><thumbnail>$$Tsyndetics_thumb_exl</thumbnail><linktopdf>$$Uhttps://www.ncbi.nlm.nih.gov/pmc/articles/PMC4471098/pdf/$$EPDF$$P50$$Gpubmedcentral$$Hfree_for_read</linktopdf><linktohtml>$$Uhttps://www.ncbi.nlm.nih.gov/pmc/articles/PMC4471098/$$EHTML$$P50$$Gpubmedcentral$$Hfree_for_read</linktohtml><link.rule.ids>230,314,727,780,784,864,885,2101,2927,23865,27923,27924,53790,53792,79471,79472</link.rule.ids><backlink>$$Uhttps://www.ncbi.nlm.nih.gov/pubmed/26083598$$D View this record in MEDLINE/PubMed$$Hfree_for_read</backlink></links><search><contributor>Ohya, Yoshikazu</contributor><creatorcontrib>Chardwiriyapreecha, Soracom</creatorcontrib><creatorcontrib>Manabe, Kunio</creatorcontrib><creatorcontrib>Iwaki, Tomoko</creatorcontrib><creatorcontrib>Kawano-Kawada, Miyuki</creatorcontrib><creatorcontrib>Sekito, Takayuki</creatorcontrib><creatorcontrib>Lunprom, Siriporn</creatorcontrib><creatorcontrib>Akiyama, Koichi</creatorcontrib><creatorcontrib>Takegawa, Kaoru</creatorcontrib><creatorcontrib>Kakinuma, Yoshimi</creatorcontrib><title>Functional Expression and Characterization of Schizosaccharomyces pombe Avt3p as a Vacuolar Amino Acid Exporter in Saccharomyces cerevisiae</title><title>PloS one</title><addtitle>PLoS One</addtitle><description>In Saccharomyces cerevisiae, Avt3p and Avt4p mediate the extrusion of several amino acids from the vacuolar lumen into the cytosol. 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These activities were inhibited by the addition of a V-ATPase inhibitor, concanamycin A, thereby suggesting that the activity of SpAvt3p is dependent on a proton electrochemical gradient generated by the action of V-ATPase. In addition, the amounts of various amino acids in the vacuolar pools of S. cerevisiae cells were decreased by the overproduction of SpAvt3p, which indicated that SpAvt3p was functional in S. cerevisiae cells. Thus, SpAvt3p is a vacuolar transporter that is involved in the export of amino acids from S. pombe vacuoles.</description><subject>Adenosine triphosphatase</subject><subject>Adenosine Triphosphatases - metabolism</subject><subject>Alanine</subject><subject>Amino Acid Sequence</subject><subject>Amino Acid Transport Systems - genetics</subject><subject>Amino Acid Transport Systems - metabolism</subject><subject>Amino acids</subject><subject>Amino Acids - metabolism</subject><subject>ATPases</subject><subject>Biological Transport</subject><subject>Concanamycin A</subject><subject>Corrosion inhibitors</subject><subject>Cytosol</subject><subject>Electrochemistry</subject><subject>Eukaryotes</subject><subject>Exports</subject><subject>Extrusion</subject><subject>Genes</subject><subject>Genetics</subject><subject>H+-transporting ATPase</subject><subject>Homology</subject><subject>Immunoblotting</subject><subject>Intracellular Membranes</subject><subject>Laboratories</subject><subject>Membrane vesicles</subject><subject>Molecular Sequence Data</subject><subject>Physiology</subject><subject>Proteins</subject><subject>Saccharomyces cerevisiae</subject><subject>Saccharomyces cerevisiae - genetics</subject><subject>Saccharomyces cerevisiae - growth & development</subject><subject>Saccharomyces cerevisiae - metabolism</subject><subject>Schizosaccharomyces - metabolism</subject><subject>Schizosaccharomyces pombe</subject><subject>Schizosaccharomyces pombe Proteins - genetics</subject><subject>Schizosaccharomyces pombe Proteins - metabolism</subject><subject>Sequence Homology, Amino Acid</subject><subject>Tyrosine</subject><subject>Vacuoles</subject><subject>Vacuoles - metabolism</subject><subject>Yeast</subject><issn>1932-6203</issn><issn>1932-6203</issn><fulltext>true</fulltext><rsrctype>article</rsrctype><creationdate>2015</creationdate><recordtype>article</recordtype><sourceid>EIF</sourceid><sourceid>ABUWG</sourceid><sourceid>AFKRA</sourceid><sourceid>AZQEC</sourceid><sourceid>BENPR</sourceid><sourceid>CCPQU</sourceid><sourceid>DWQXO</sourceid><sourceid>GNUQQ</sourceid><sourceid>DOA</sourceid><recordid>eNqNk9Fu0zAUhiMEYmPwBggsISG4aLFjJ3FukKpqg0qTJlHYreXYx62rJO7spNr2Crw0Du2mBu0C-cL28ff_to99kuQtwVNCC_Jl43rfynq6dS1MMaE4Y-mz5JSUNJ3kKabPj8YnyasQNhhnlOf5y-QkzTGnWclPk98Xfas666ITOr_degghTpBsNZqvpZeqA2_v5UAgZ9BSre29C1KpuOiaOwUBbV1TAZrtOrpFMiCJrqXqXS09mjW2dWimrB68nY9eyLZoOZIr8LCzwUp4nbwwsg7w5tCfJb8uzn_Ov08ur74t5rPLiSoy3k24qdLCgOTSlMrI0uScVNzkrCpIplNmWJWlUILRioHGNKs0aKAm4xnGhFN6lrzf-25rF8Qhj0GQnJd5ykmeR2KxJ7STG7H1tpH-Tjhpxd-A8yshfWdVDYJpk-dFpgjDhlFKOS20ZmkMMl5hkkWvr4fd-qoBraDtvKxHpuOV1q7Fyu0EYwXBJY8Gnw4G3t30EDrR2KCgrmULrh_OXeKUpITjiH74B336dgdqJeMFbGtc3FcNpmLGYoJwPDqL1PQJKjYNjVXx0xkb4yPB55EgMh3cdivZhyAWyx__z15dj9mPR-waZN2tg6v74U-GMcj2oPIuBA_mMckEi6FmHrIhhpoRh5qJsnfHD_QoeigS-gch2xOy</recordid><startdate>20150617</startdate><enddate>20150617</enddate><creator>Chardwiriyapreecha, Soracom</creator><creator>Manabe, Kunio</creator><creator>Iwaki, Tomoko</creator><creator>Kawano-Kawada, Miyuki</creator><creator>Sekito, Takayuki</creator><creator>Lunprom, Siriporn</creator><creator>Akiyama, Koichi</creator><creator>Takegawa, Kaoru</creator><creator>Kakinuma, Yoshimi</creator><general>Public Library of Science</general><general>Public Library of Science (PLoS)</general><scope>CGR</scope><scope>CUY</scope><scope>CVF</scope><scope>ECM</scope><scope>EIF</scope><scope>NPM</scope><scope>AAYXX</scope><scope>CITATION</scope><scope>IOV</scope><scope>ISR</scope><scope>3V.</scope><scope>7QG</scope><scope>7QL</scope><scope>7QO</scope><scope>7RV</scope><scope>7SN</scope><scope>7SS</scope><scope>7T5</scope><scope>7TG</scope><scope>7TM</scope><scope>7U9</scope><scope>7X2</scope><scope>7X7</scope><scope>7XB</scope><scope>88E</scope><scope>8AO</scope><scope>8C1</scope><scope>8FD</scope><scope>8FE</scope><scope>8FG</scope><scope>8FH</scope><scope>8FI</scope><scope>8FJ</scope><scope>8FK</scope><scope>ABJCF</scope><scope>ABUWG</scope><scope>AEUYN</scope><scope>AFKRA</scope><scope>ARAPS</scope><scope>ATCPS</scope><scope>AZQEC</scope><scope>BBNVY</scope><scope>BENPR</scope><scope>BGLVJ</scope><scope>BHPHI</scope><scope>C1K</scope><scope>CCPQU</scope><scope>D1I</scope><scope>DWQXO</scope><scope>FR3</scope><scope>FYUFA</scope><scope>GHDGH</scope><scope>GNUQQ</scope><scope>H94</scope><scope>HCIFZ</scope><scope>K9.</scope><scope>KB.</scope><scope>KB0</scope><scope>KL.</scope><scope>L6V</scope><scope>LK8</scope><scope>M0K</scope><scope>M0S</scope><scope>M1P</scope><scope>M7N</scope><scope>M7P</scope><scope>M7S</scope><scope>NAPCQ</scope><scope>P5Z</scope><scope>P62</scope><scope>P64</scope><scope>PATMY</scope><scope>PDBOC</scope><scope>PIMPY</scope><scope>PQEST</scope><scope>PQQKQ</scope><scope>PQUKI</scope><scope>PRINS</scope><scope>PTHSS</scope><scope>PYCSY</scope><scope>RC3</scope><scope>7X8</scope><scope>5PM</scope><scope>DOA</scope></search><sort><creationdate>20150617</creationdate><title>Functional Expression and Characterization of Schizosaccharomyces pombe Avt3p as a Vacuolar Amino Acid Exporter in Saccharomyces cerevisiae</title><author>Chardwiriyapreecha, Soracom ; Manabe, Kunio ; Iwaki, Tomoko ; Kawano-Kawada, Miyuki ; Sekito, Takayuki ; Lunprom, Siriporn ; Akiyama, Koichi ; Takegawa, Kaoru ; Kakinuma, Yoshimi</author></sort><facets><frbrtype>5</frbrtype><frbrgroupid>cdi_FETCH-LOGICAL-c758t-8fb27fea8af9cfa9f681b8f64b715d24f4b52e9efdc4ed035bdede3f585001833</frbrgroupid><rsrctype>articles</rsrctype><prefilter>articles</prefilter><language>eng</language><creationdate>2015</creationdate><topic>Adenosine triphosphatase</topic><topic>Adenosine Triphosphatases - 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Academic</collection><collection>PubMed Central (Full Participant titles)</collection><collection>DOAJ Directory of Open Access Journals</collection><jtitle>PloS one</jtitle></facets><delivery><delcategory>Remote Search Resource</delcategory><fulltext>fulltext</fulltext></delivery><addata><au>Chardwiriyapreecha, Soracom</au><au>Manabe, Kunio</au><au>Iwaki, Tomoko</au><au>Kawano-Kawada, Miyuki</au><au>Sekito, Takayuki</au><au>Lunprom, Siriporn</au><au>Akiyama, Koichi</au><au>Takegawa, Kaoru</au><au>Kakinuma, Yoshimi</au><au>Ohya, Yoshikazu</au><format>journal</format><genre>article</genre><ristype>JOUR</ristype><atitle>Functional Expression and Characterization of Schizosaccharomyces pombe Avt3p as a Vacuolar Amino Acid Exporter in Saccharomyces cerevisiae</atitle><jtitle>PloS one</jtitle><addtitle>PLoS One</addtitle><date>2015-06-17</date><risdate>2015</risdate><volume>10</volume><issue>6</issue><spage>e0130542</spage><epage>e0130542</epage><pages>e0130542-e0130542</pages><issn>1932-6203</issn><eissn>1932-6203</eissn><abstract>In Saccharomyces cerevisiae, Avt3p and Avt4p mediate the extrusion of several amino acids from the vacuolar lumen into the cytosol. SpAvt3p of Schizosaccharomyces pombe, a homologue of these vacuolar amino acid transporters, has been indicated to be involved in spore formation. In this study, we confirmed that GFP-SpAvt3p localized to the vacuolar membrane in S. pombe. The amounts of various amino acids increased significantly in the vacuolar pool of avt3Δ cells, but decreased in that of avt3+-overexpressing avt3Δ cells. These results suggest that SpAvt3p participates in the vacuolar compartmentalization of amino acids in S. pombe. To examine the export activity of SpAvt3p, we expressed the avt3+ gene in S. cerevisiae cells. We found that the heterologously overproduced GFP-SpAvt3p localized to the vacuolar membrane in S. cerevisiae. Using the vacuolar membrane vesicles isolated from avt3+-overexpressing S. cerevisiae cells, we detected the export activities of alanine and tyrosine in an ATP-dependent manner. These activities were inhibited by the addition of a V-ATPase inhibitor, concanamycin A, thereby suggesting that the activity of SpAvt3p is dependent on a proton electrochemical gradient generated by the action of V-ATPase. In addition, the amounts of various amino acids in the vacuolar pools of S. cerevisiae cells were decreased by the overproduction of SpAvt3p, which indicated that SpAvt3p was functional in S. cerevisiae cells. Thus, SpAvt3p is a vacuolar transporter that is involved in the export of amino acids from S. pombe vacuoles.</abstract><cop>United States</cop><pub>Public Library of Science</pub><pmid>26083598</pmid><doi>10.1371/journal.pone.0130542</doi><oa>free_for_read</oa></addata></record> |
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| language | eng |
| recordid | cdi_plos_journals_1689628166 |
| source | MEDLINE; DOAJ Directory of Open Access Journals; Public Library of Science (PLoS); EZB-FREE-00999 freely available EZB journals; PubMed Central; Free Full-Text Journals in Chemistry |
| subjects | Adenosine triphosphatase Adenosine Triphosphatases - metabolism Alanine Amino Acid Sequence Amino Acid Transport Systems - genetics Amino Acid Transport Systems - metabolism Amino acids Amino Acids - metabolism ATPases Biological Transport Concanamycin A Corrosion inhibitors Cytosol Electrochemistry Eukaryotes Exports Extrusion Genes Genetics H+-transporting ATPase Homology Immunoblotting Intracellular Membranes Laboratories Membrane vesicles Molecular Sequence Data Physiology Proteins Saccharomyces cerevisiae Saccharomyces cerevisiae - genetics Saccharomyces cerevisiae - growth & development Saccharomyces cerevisiae - metabolism Schizosaccharomyces - metabolism Schizosaccharomyces pombe Schizosaccharomyces pombe Proteins - genetics Schizosaccharomyces pombe Proteins - metabolism Sequence Homology, Amino Acid Tyrosine Vacuoles Vacuoles - metabolism Yeast |
| title | Functional Expression and Characterization of Schizosaccharomyces pombe Avt3p as a Vacuolar Amino Acid Exporter in Saccharomyces cerevisiae |
| url | https://sfx.bib-bvb.de/sfx_tum?ctx_ver=Z39.88-2004&ctx_enc=info:ofi/enc:UTF-8&ctx_tim=2025-01-08T23%3A26%3A17IST&url_ver=Z39.88-2004&url_ctx_fmt=infofi/fmt:kev:mtx:ctx&rfr_id=info:sid/primo.exlibrisgroup.com:primo3-Article-gale_plos_&rft_val_fmt=info:ofi/fmt:kev:mtx:journal&rft.genre=article&rft.atitle=Functional%20Expression%20and%20Characterization%20of%20Schizosaccharomyces%20pombe%20Avt3p%20as%20a%20Vacuolar%20Amino%20Acid%20Exporter%20in%20Saccharomyces%20cerevisiae&rft.jtitle=PloS%20one&rft.au=Chardwiriyapreecha,%20Soracom&rft.date=2015-06-17&rft.volume=10&rft.issue=6&rft.spage=e0130542&rft.epage=e0130542&rft.pages=e0130542-e0130542&rft.issn=1932-6203&rft.eissn=1932-6203&rft_id=info:doi/10.1371/journal.pone.0130542&rft_dat=%3Cgale_plos_%3EA418306484%3C/gale_plos_%3E%3Curl%3E%3C/url%3E&disable_directlink=true&sfx.directlink=off&sfx.report_link=0&rft_id=info:oai/&rft_pqid=1689628166&rft_id=info:pmid/26083598&rft_galeid=A418306484&rft_doaj_id=oai_doaj_org_article_4df6675c140f4333837dd42f6648b015&rfr_iscdi=true |