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<title xml:lang="en">Evidence for annexin A6-dependent plasma membrane remodelling of lipid domains</title>
<author>
<name sortKey="Alvarez Guaita, Anna" sort="Alvarez Guaita, Anna" uniqKey="Alvarez Guaita A" first="Anna" last="Alvarez-Guaita">Anna Alvarez-Guaita</name>
<affiliation>
<nlm:aff id="au1">
<institution>Departament de Biologia Cel·lular, Immunologia i Neurociències, Facultat de Medicina, Universitat de Barcelona</institution>
<addr-line>Barcelona, Spain</addr-line>
</nlm:aff>
</affiliation>
</author>
<author>
<name sortKey="Vila De Muga, Sandra" sort="Vila De Muga, Sandra" uniqKey="Vila De Muga S" first="Sandra" last="Vilà De Muga">Sandra Vilà De Muga</name>
<affiliation>
<nlm:aff id="au1">
<institution>Departament de Biologia Cel·lular, Immunologia i Neurociències, Facultat de Medicina, Universitat de Barcelona</institution>
<addr-line>Barcelona, Spain</addr-line>
</nlm:aff>
</affiliation>
</author>
<author>
<name sortKey="Owen, Dylan M" sort="Owen, Dylan M" uniqKey="Owen D" first="Dylan M" last="Owen">Dylan M. Owen</name>
<affiliation>
<nlm:aff id="au2">
<institution>Center for Vascular Research, The University of New South Wales</institution>
<addr-line>Sydney, NSW, Australia</addr-line>
</nlm:aff>
</affiliation>
</author>
<author>
<name sortKey="Williamson, David" sort="Williamson, David" uniqKey="Williamson D" first="David" last="Williamson">David Williamson</name>
<affiliation>
<nlm:aff id="au2">
<institution>Center for Vascular Research, The University of New South Wales</institution>
<addr-line>Sydney, NSW, Australia</addr-line>
</nlm:aff>
</affiliation>
</author>
<author>
<name sortKey="Magenau, Astrid" sort="Magenau, Astrid" uniqKey="Magenau A" first="Astrid" last="Magenau">Astrid Magenau</name>
<affiliation>
<nlm:aff id="au2">
<institution>Center for Vascular Research, The University of New South Wales</institution>
<addr-line>Sydney, NSW, Australia</addr-line>
</nlm:aff>
</affiliation>
</author>
<author>
<name sortKey="Garcia Melero, Ana" sort="Garcia Melero, Ana" uniqKey="Garcia Melero A" first="Ana" last="García-Melero">Ana García-Melero</name>
<affiliation>
<nlm:aff id="au1">
<institution>Departament de Biologia Cel·lular, Immunologia i Neurociències, Facultat de Medicina, Universitat de Barcelona</institution>
<addr-line>Barcelona, Spain</addr-line>
</nlm:aff>
</affiliation>
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<author>
<name sortKey="Reverter, Meritxell" sort="Reverter, Meritxell" uniqKey="Reverter M" first="Meritxell" last="Reverter">Meritxell Reverter</name>
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<institution>Departament de Biologia Cel·lular, Immunologia i Neurociències, Facultat de Medicina, Universitat de Barcelona</institution>
<addr-line>Barcelona, Spain</addr-line>
</nlm:aff>
</affiliation>
</author>
<author>
<name sortKey="Hoque, Monira" sort="Hoque, Monira" uniqKey="Hoque M" first="Monira" last="Hoque">Monira Hoque</name>
<affiliation>
<nlm:aff id="au3">
<institution>Faculty of Pharmacy, University of Sydney</institution>
<addr-line>Sydney, NSW, Australia</addr-line>
</nlm:aff>
</affiliation>
</author>
<author>
<name sortKey="Cairns, Rose" sort="Cairns, Rose" uniqKey="Cairns R" first="Rose" last="Cairns">Rose Cairns</name>
<affiliation>
<nlm:aff id="au3">
<institution>Faculty of Pharmacy, University of Sydney</institution>
<addr-line>Sydney, NSW, Australia</addr-line>
</nlm:aff>
</affiliation>
</author>
<author>
<name sortKey="Cornely, Rhea" sort="Cornely, Rhea" uniqKey="Cornely R" first="Rhea" last="Cornely">Rhea Cornely</name>
<affiliation>
<nlm:aff id="au2">
<institution>Center for Vascular Research, The University of New South Wales</institution>
<addr-line>Sydney, NSW, Australia</addr-line>
</nlm:aff>
</affiliation>
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<author>
<name sortKey="Tebar, Francesc" sort="Tebar, Francesc" uniqKey="Tebar F" first="Francesc" last="Tebar">Francesc Tebar</name>
<affiliation>
<nlm:aff id="au1">
<institution>Departament de Biologia Cel·lular, Immunologia i Neurociències, Facultat de Medicina, Universitat de Barcelona</institution>
<addr-line>Barcelona, Spain</addr-line>
</nlm:aff>
</affiliation>
</author>
<author>
<name sortKey="Grewal, Thomas" sort="Grewal, Thomas" uniqKey="Grewal T" first="Thomas" last="Grewal">Thomas Grewal</name>
<affiliation>
<nlm:aff id="au3">
<institution>Faculty of Pharmacy, University of Sydney</institution>
<addr-line>Sydney, NSW, Australia</addr-line>
</nlm:aff>
</affiliation>
</author>
<author>
<name sortKey="Gaus, Katharina" sort="Gaus, Katharina" uniqKey="Gaus K" first="Katharina" last="Gaus">Katharina Gaus</name>
<affiliation>
<nlm:aff id="au2">
<institution>Center for Vascular Research, The University of New South Wales</institution>
<addr-line>Sydney, NSW, Australia</addr-line>
</nlm:aff>
</affiliation>
</author>
<author>
<name sortKey="Ayala Sanmartin, Jesus" sort="Ayala Sanmartin, Jesus" uniqKey="Ayala Sanmartin J" first="Jesús" last="Ayala-Sanmartín">Jesús Ayala-Sanmartín</name>
<affiliation>
<nlm:aff id="au4">
<institution>Centre National de la Recherche Scientifique (CNRS), École Normale Supérieure (ENS) and Université Pierre et Marie Curie (UPMC)</institution>
<addr-line>Paris, France</addr-line>
</nlm:aff>
</affiliation>
</author>
<author>
<name sortKey="Enrich, Carlos" sort="Enrich, Carlos" uniqKey="Enrich C" first="Carlos" last="Enrich">Carlos Enrich</name>
<affiliation>
<nlm:aff id="au1">
<institution>Departament de Biologia Cel·lular, Immunologia i Neurociències, Facultat de Medicina, Universitat de Barcelona</institution>
<addr-line>Barcelona, Spain</addr-line>
</nlm:aff>
</affiliation>
<affiliation>
<nlm:aff id="au5">
<institution>Centre de Recerca Biomèdica CELLEX, Institut d'Investigacions Biomèdiques August Pi i Sunyer (IDIBAPS)</institution>
<addr-line>Barcelona, Spain</addr-line>
</nlm:aff>
</affiliation>
</author>
<author>
<name sortKey="Rentero, Carles" sort="Rentero, Carles" uniqKey="Rentero C" first="Carles" last="Rentero">Carles Rentero</name>
<affiliation>
<nlm:aff id="au1">
<institution>Departament de Biologia Cel·lular, Immunologia i Neurociències, Facultat de Medicina, Universitat de Barcelona</institution>
<addr-line>Barcelona, Spain</addr-line>
</nlm:aff>
</affiliation>
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<idno type="doi">10.1111/bph.13022</idno>
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<title xml:lang="en" level="a" type="main">Evidence for annexin A6-dependent plasma membrane remodelling of lipid domains</title>
<author>
<name sortKey="Alvarez Guaita, Anna" sort="Alvarez Guaita, Anna" uniqKey="Alvarez Guaita A" first="Anna" last="Alvarez-Guaita">Anna Alvarez-Guaita</name>
<affiliation>
<nlm:aff id="au1">
<institution>Departament de Biologia Cel·lular, Immunologia i Neurociències, Facultat de Medicina, Universitat de Barcelona</institution>
<addr-line>Barcelona, Spain</addr-line>
</nlm:aff>
</affiliation>
</author>
<author>
<name sortKey="Vila De Muga, Sandra" sort="Vila De Muga, Sandra" uniqKey="Vila De Muga S" first="Sandra" last="Vilà De Muga">Sandra Vilà De Muga</name>
<affiliation>
<nlm:aff id="au1">
<institution>Departament de Biologia Cel·lular, Immunologia i Neurociències, Facultat de Medicina, Universitat de Barcelona</institution>
<addr-line>Barcelona, Spain</addr-line>
</nlm:aff>
</affiliation>
</author>
<author>
<name sortKey="Owen, Dylan M" sort="Owen, Dylan M" uniqKey="Owen D" first="Dylan M" last="Owen">Dylan M. Owen</name>
<affiliation>
<nlm:aff id="au2">
<institution>Center for Vascular Research, The University of New South Wales</institution>
<addr-line>Sydney, NSW, Australia</addr-line>
</nlm:aff>
</affiliation>
</author>
<author>
<name sortKey="Williamson, David" sort="Williamson, David" uniqKey="Williamson D" first="David" last="Williamson">David Williamson</name>
<affiliation>
<nlm:aff id="au2">
<institution>Center for Vascular Research, The University of New South Wales</institution>
<addr-line>Sydney, NSW, Australia</addr-line>
</nlm:aff>
</affiliation>
</author>
<author>
<name sortKey="Magenau, Astrid" sort="Magenau, Astrid" uniqKey="Magenau A" first="Astrid" last="Magenau">Astrid Magenau</name>
<affiliation>
<nlm:aff id="au2">
<institution>Center for Vascular Research, The University of New South Wales</institution>
<addr-line>Sydney, NSW, Australia</addr-line>
</nlm:aff>
</affiliation>
</author>
<author>
<name sortKey="Garcia Melero, Ana" sort="Garcia Melero, Ana" uniqKey="Garcia Melero A" first="Ana" last="García-Melero">Ana García-Melero</name>
<affiliation>
<nlm:aff id="au1">
<institution>Departament de Biologia Cel·lular, Immunologia i Neurociències, Facultat de Medicina, Universitat de Barcelona</institution>
<addr-line>Barcelona, Spain</addr-line>
</nlm:aff>
</affiliation>
</author>
<author>
<name sortKey="Reverter, Meritxell" sort="Reverter, Meritxell" uniqKey="Reverter M" first="Meritxell" last="Reverter">Meritxell Reverter</name>
<affiliation>
<nlm:aff id="au1">
<institution>Departament de Biologia Cel·lular, Immunologia i Neurociències, Facultat de Medicina, Universitat de Barcelona</institution>
<addr-line>Barcelona, Spain</addr-line>
</nlm:aff>
</affiliation>
</author>
<author>
<name sortKey="Hoque, Monira" sort="Hoque, Monira" uniqKey="Hoque M" first="Monira" last="Hoque">Monira Hoque</name>
<affiliation>
<nlm:aff id="au3">
<institution>Faculty of Pharmacy, University of Sydney</institution>
<addr-line>Sydney, NSW, Australia</addr-line>
</nlm:aff>
</affiliation>
</author>
<author>
<name sortKey="Cairns, Rose" sort="Cairns, Rose" uniqKey="Cairns R" first="Rose" last="Cairns">Rose Cairns</name>
<affiliation>
<nlm:aff id="au3">
<institution>Faculty of Pharmacy, University of Sydney</institution>
<addr-line>Sydney, NSW, Australia</addr-line>
</nlm:aff>
</affiliation>
</author>
<author>
<name sortKey="Cornely, Rhea" sort="Cornely, Rhea" uniqKey="Cornely R" first="Rhea" last="Cornely">Rhea Cornely</name>
<affiliation>
<nlm:aff id="au2">
<institution>Center for Vascular Research, The University of New South Wales</institution>
<addr-line>Sydney, NSW, Australia</addr-line>
</nlm:aff>
</affiliation>
</author>
<author>
<name sortKey="Tebar, Francesc" sort="Tebar, Francesc" uniqKey="Tebar F" first="Francesc" last="Tebar">Francesc Tebar</name>
<affiliation>
<nlm:aff id="au1">
<institution>Departament de Biologia Cel·lular, Immunologia i Neurociències, Facultat de Medicina, Universitat de Barcelona</institution>
<addr-line>Barcelona, Spain</addr-line>
</nlm:aff>
</affiliation>
</author>
<author>
<name sortKey="Grewal, Thomas" sort="Grewal, Thomas" uniqKey="Grewal T" first="Thomas" last="Grewal">Thomas Grewal</name>
<affiliation>
<nlm:aff id="au3">
<institution>Faculty of Pharmacy, University of Sydney</institution>
<addr-line>Sydney, NSW, Australia</addr-line>
</nlm:aff>
</affiliation>
</author>
<author>
<name sortKey="Gaus, Katharina" sort="Gaus, Katharina" uniqKey="Gaus K" first="Katharina" last="Gaus">Katharina Gaus</name>
<affiliation>
<nlm:aff id="au2">
<institution>Center for Vascular Research, The University of New South Wales</institution>
<addr-line>Sydney, NSW, Australia</addr-line>
</nlm:aff>
</affiliation>
</author>
<author>
<name sortKey="Ayala Sanmartin, Jesus" sort="Ayala Sanmartin, Jesus" uniqKey="Ayala Sanmartin J" first="Jesús" last="Ayala-Sanmartín">Jesús Ayala-Sanmartín</name>
<affiliation>
<nlm:aff id="au4">
<institution>Centre National de la Recherche Scientifique (CNRS), École Normale Supérieure (ENS) and Université Pierre et Marie Curie (UPMC)</institution>
<addr-line>Paris, France</addr-line>
</nlm:aff>
</affiliation>
</author>
<author>
<name sortKey="Enrich, Carlos" sort="Enrich, Carlos" uniqKey="Enrich C" first="Carlos" last="Enrich">Carlos Enrich</name>
<affiliation>
<nlm:aff id="au1">
<institution>Departament de Biologia Cel·lular, Immunologia i Neurociències, Facultat de Medicina, Universitat de Barcelona</institution>
<addr-line>Barcelona, Spain</addr-line>
</nlm:aff>
</affiliation>
<affiliation>
<nlm:aff id="au5">
<institution>Centre de Recerca Biomèdica CELLEX, Institut d'Investigacions Biomèdiques August Pi i Sunyer (IDIBAPS)</institution>
<addr-line>Barcelona, Spain</addr-line>
</nlm:aff>
</affiliation>
</author>
<author>
<name sortKey="Rentero, Carles" sort="Rentero, Carles" uniqKey="Rentero C" first="Carles" last="Rentero">Carles Rentero</name>
<affiliation>
<nlm:aff id="au1">
<institution>Departament de Biologia Cel·lular, Immunologia i Neurociències, Facultat de Medicina, Universitat de Barcelona</institution>
<addr-line>Barcelona, Spain</addr-line>
</nlm:aff>
</affiliation>
</author>
</analytic>
<series>
<title level="j">British Journal of Pharmacology</title>
<idno type="ISSN">0007-1188</idno>
<idno type="eISSN">1476-5381</idno>
<imprint>
<date when="2015">2015</date>
</imprint>
</series>
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<front>
<div type="abstract" xml:lang="en">
<sec>
<title>Background and Purpose</title>
<p>Annexin A6 (AnxA6) is a calcium-dependent phospholipid-binding protein that can be recruited to the plasma membrane to function as a scaffolding protein to regulate signal complex formation, endo- and exocytic pathways as well as distribution of cellular cholesterol. Here, we have investigated how AnxA6 influences the membrane order.</p>
</sec>
<sec>
<title>Experimental Approach</title>
<p>We used Laurdan and di-4-ANEPPDHQ staining in (i) artificial membranes; (ii) live cells to investigate membrane packing and ordered lipid phases; and (iii) a super-resolution imaging (photoactivated localization microscopy, PALM) and Ripley's K second-order point pattern analysis approach to assess how AnxA6 regulates plasma membrane order domains and protein clustering.</p>
</sec>
<sec>
<title>Key Results</title>
<p>In artificial membranes, purified AnxA6 induced a global increase in membrane order. However, confocal microscopy using di-4-ANEPPDHQ in live cells showed that cells expressing AnxA6, which reduces plasma membrane cholesterol levels and modifies the actin cytoskeleton meshwork, displayed a decrease in membrane order (∼15 and 30% in A431 and MEF cells respectively). PALM data from Lck10 and Src15 membrane raft/non-raft markers revealed that AnxA6 expression induced clustering of both raft and non-raft markers. Altered clustering of Lck10 and Src15 in cells expressing AnxA6 was also observed after cholesterol extraction with methyl-β-cyclodextrin or actin cytoskeleton disruption with latrunculin B.</p>
</sec>
<sec>
<title>Conclusions and Implications</title>
<p>AnxA6-induced plasma membrane remodelling indicated that elevated AnxA6 expression decreased membrane order through the regulation of cellular cholesterol homeostasis and the actin cytoskeleton. This study provides the first evidence from live cells that support current models of annexins as membrane organizers.</p>
</sec>
</div>
</front>
</TEI>
<pmc article-type="research-article">
<pmc-comment>The publisher of this article does not allow downloading of the full text in XML form.</pmc-comment>
<front>
<journal-meta>
<journal-id journal-id-type="nlm-ta">Br J Pharmacol</journal-id>
<journal-id journal-id-type="iso-abbrev">Br. J. Pharmacol</journal-id>
<journal-id journal-id-type="publisher-id">bph</journal-id>
<journal-title-group>
<journal-title>British Journal of Pharmacology</journal-title>
</journal-title-group>
<issn pub-type="ppub">0007-1188</issn>
<issn pub-type="epub">1476-5381</issn>
<publisher>
<publisher-name>BlackWell Publishing Ltd</publisher-name>
<publisher-loc>Oxford, UK</publisher-loc>
</publisher>
</journal-meta>
<article-meta>
<article-id pub-id-type="pmid">25409976</article-id>
<article-id pub-id-type="pmc">4376448</article-id>
<article-id pub-id-type="doi">10.1111/bph.13022</article-id>
<article-categories>
<subj-group subj-group-type="heading">
<subject>Themed Section: Annexins Vii Programme</subject>
<subj-group subj-group-type="heading">
<subject>Research Paper</subject>
</subj-group>
</subj-group>
</article-categories>
<title-group>
<article-title>Evidence for annexin A6-dependent plasma membrane remodelling of lipid domains</article-title>
</title-group>
<contrib-group>
<contrib contrib-type="author">
<name>
<surname>Alvarez-Guaita</surname>
<given-names>Anna</given-names>
</name>
<xref ref-type="aff" rid="au1">1</xref>
<xref ref-type="author-notes" rid="fn1">*</xref>
</contrib>
<contrib contrib-type="author">
<name>
<surname>Vilà de Muga</surname>
<given-names>Sandra</given-names>
</name>
<xref ref-type="aff" rid="au1">1</xref>
<xref ref-type="author-notes" rid="fn1">*</xref>
</contrib>
<contrib contrib-type="author">
<name>
<surname>Owen</surname>
<given-names>Dylan M</given-names>
</name>
<xref ref-type="aff" rid="au2">2</xref>
<xref ref-type="author-notes" rid="fn2"></xref>
</contrib>
<contrib contrib-type="author">
<name>
<surname>Williamson</surname>
<given-names>David</given-names>
</name>
<xref ref-type="aff" rid="au2">2</xref>
<xref ref-type="author-notes" rid="fn3"></xref>
</contrib>
<contrib contrib-type="author">
<name>
<surname>Magenau</surname>
<given-names>Astrid</given-names>
</name>
<xref ref-type="aff" rid="au2">2</xref>
<xref ref-type="author-notes" rid="fn4">§</xref>
</contrib>
<contrib contrib-type="author">
<name>
<surname>García-Melero</surname>
<given-names>Ana</given-names>
</name>
<xref ref-type="aff" rid="au1">1</xref>
</contrib>
<contrib contrib-type="author">
<name>
<surname>Reverter</surname>
<given-names>Meritxell</given-names>
</name>
<xref ref-type="aff" rid="au1">1</xref>
</contrib>
<contrib contrib-type="author">
<name>
<surname>Hoque</surname>
<given-names>Monira</given-names>
</name>
<xref ref-type="aff" rid="au3">3</xref>
</contrib>
<contrib contrib-type="author">
<name>
<surname>Cairns</surname>
<given-names>Rose</given-names>
</name>
<xref ref-type="aff" rid="au3">3</xref>
</contrib>
<contrib contrib-type="author">
<name>
<surname>Cornely</surname>
<given-names>Rhea</given-names>
</name>
<xref ref-type="aff" rid="au2">2</xref>
</contrib>
<contrib contrib-type="author">
<name>
<surname>Tebar</surname>
<given-names>Francesc</given-names>
</name>
<xref ref-type="aff" rid="au1">1</xref>
</contrib>
<contrib contrib-type="author">
<name>
<surname>Grewal</surname>
<given-names>Thomas</given-names>
</name>
<xref ref-type="aff" rid="au3">3</xref>
</contrib>
<contrib contrib-type="author">
<name>
<surname>Gaus</surname>
<given-names>Katharina</given-names>
</name>
<xref ref-type="aff" rid="au2">2</xref>
</contrib>
<contrib contrib-type="author">
<name>
<surname>Ayala-Sanmartín</surname>
<given-names>Jesús</given-names>
</name>
<xref ref-type="aff" rid="au4">4</xref>
</contrib>
<contrib contrib-type="author">
<name>
<surname>Enrich</surname>
<given-names>Carlos</given-names>
</name>
<xref ref-type="aff" rid="au1">1</xref>
<xref ref-type="aff" rid="au5">5</xref>
<xref ref-type="corresp" rid="cor1"></xref>
</contrib>
<contrib contrib-type="author">
<name>
<surname>Rentero</surname>
<given-names>Carles</given-names>
</name>
<xref ref-type="aff" rid="au1">1</xref>
<xref ref-type="corresp" rid="cor1"></xref>
</contrib>
<aff id="au1">
<label>1</label>
<institution>Departament de Biologia Cel·lular, Immunologia i Neurociències, Facultat de Medicina, Universitat de Barcelona</institution>
<addr-line>Barcelona, Spain</addr-line>
</aff>
<aff id="au2">
<label>2</label>
<institution>Center for Vascular Research, The University of New South Wales</institution>
<addr-line>Sydney, NSW, Australia</addr-line>
</aff>
<aff id="au3">
<label>3</label>
<institution>Faculty of Pharmacy, University of Sydney</institution>
<addr-line>Sydney, NSW, Australia</addr-line>
</aff>
<aff id="au4">
<label>4</label>
<institution>Centre National de la Recherche Scientifique (CNRS), École Normale Supérieure (ENS) and Université Pierre et Marie Curie (UPMC)</institution>
<addr-line>Paris, France</addr-line>
</aff>
<aff id="au5">
<label>5</label>
<institution>Centre de Recerca Biomèdica CELLEX, Institut d'Investigacions Biomèdiques August Pi i Sunyer (IDIBAPS)</institution>
<addr-line>Barcelona, Spain</addr-line>
</aff>
</contrib-group>
<author-notes>
<corresp id="cor1">
<bold>Correspondence</bold>
, Carles Rentero and Carlos Enrich, Departament de Biologia Cel·lular, Immunologia i Neurociències, Facultat de Medicina, Universitat de Barcelona, 08036 Barcelona, Spain. E-mail:
<email>carles.rentero@ub.edu</email>
;
<email>enrich@ub.edu</email>
</corresp>
<fn id="fn1">
<label>*</label>
<p>Both authors contributed equally.</p>
</fn>
<fn id="fn2">
<label></label>
<p>Present addresses: Department of Physics and Randall Division of Cell and Molecular Biophysics, King's College London London, WC2R 2LS, UK</p>
</fn>
<fn id="fn3">
<label></label>
<p>Faculty of Life Sciences, University of Manchester Manchester, M13 9PT, UK</p>
</fn>
<fn id="fn4">
<label>§</label>
<p>Garvan Institute of Medical Research and Kinghorn Cancer Centre, Cancer Research Program, St. Vincent's Clinical School, Faculty of Medicine, University of New South Wales Sydney, NSW, 2010, Australia</p>
</fn>
</author-notes>
<pub-date pub-type="ppub">
<month>4</month>
<year>2015</year>
</pub-date>
<pub-date pub-type="epub">
<day>20</day>
<month>1</month>
<year>2015</year>
</pub-date>
<volume>172</volume>
<issue>7</issue>
<fpage>1677</fpage>
<lpage>1690</lpage>
<history>
<date date-type="received">
<day>02</day>
<month>4</month>
<year>2014</year>
</date>
<date date-type="rev-recd">
<day>11</day>
<month>11</month>
<year>2014</year>
</date>
<date date-type="accepted">
<day>14</day>
<month>11</month>
<year>2014</year>
</date>
</history>
<permissions>
<copyright-statement>Copyright © 2015 The British Pharmacological Society</copyright-statement>
<copyright-year>2015</copyright-year>
</permissions>
<abstract>
<sec>
<title>Background and Purpose</title>
<p>Annexin A6 (AnxA6) is a calcium-dependent phospholipid-binding protein that can be recruited to the plasma membrane to function as a scaffolding protein to regulate signal complex formation, endo- and exocytic pathways as well as distribution of cellular cholesterol. Here, we have investigated how AnxA6 influences the membrane order.</p>
</sec>
<sec>
<title>Experimental Approach</title>
<p>We used Laurdan and di-4-ANEPPDHQ staining in (i) artificial membranes; (ii) live cells to investigate membrane packing and ordered lipid phases; and (iii) a super-resolution imaging (photoactivated localization microscopy, PALM) and Ripley's K second-order point pattern analysis approach to assess how AnxA6 regulates plasma membrane order domains and protein clustering.</p>
</sec>
<sec>
<title>Key Results</title>
<p>In artificial membranes, purified AnxA6 induced a global increase in membrane order. However, confocal microscopy using di-4-ANEPPDHQ in live cells showed that cells expressing AnxA6, which reduces plasma membrane cholesterol levels and modifies the actin cytoskeleton meshwork, displayed a decrease in membrane order (∼15 and 30% in A431 and MEF cells respectively). PALM data from Lck10 and Src15 membrane raft/non-raft markers revealed that AnxA6 expression induced clustering of both raft and non-raft markers. Altered clustering of Lck10 and Src15 in cells expressing AnxA6 was also observed after cholesterol extraction with methyl-β-cyclodextrin or actin cytoskeleton disruption with latrunculin B.</p>
</sec>
<sec>
<title>Conclusions and Implications</title>
<p>AnxA6-induced plasma membrane remodelling indicated that elevated AnxA6 expression decreased membrane order through the regulation of cellular cholesterol homeostasis and the actin cytoskeleton. This study provides the first evidence from live cells that support current models of annexins as membrane organizers.</p>
</sec>
</abstract>
</article-meta>
</front>
</pmc>
</record>

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