Podoplanin is a substrate of presenilin-1/γ-secretase

Podoplanin (PDPN) is a mucin-like transmembrane glycoprotein that plays an important role in development and cancer. Here, we provide evidence that the intracellular domain (ICD) of podoplanin is released into the cytosol following a sequential proteolytic processing by a metalloprotease and γ-secre...

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Detalhes bibliográficos
Autores: Yurrita, María M., Fernández-Muñoz, Beatriz, Castillo, Gaelle del, Martín-Villar, Ester, Renart, Jaime, Quintanilla, Miguel
Formato: artículo
Estado:Versión enviada para evaluación y publicación
Fecha de publicación:2014
País:España
Recursos:Consejo Superior de Investigaciones Científicas (CSIC)
Repositorio:DIGITAL.CSIC. Repositorio Institucional del CSIC
OAI Identifier:oai:digital.csic.es:10261/111808
Acesso em linha:http://hdl.handle.net/10261/111808
Access Level:acceso abierto
Palavra-chave:Metalloprotease
γ-Secretase
Podoplanin
Proteolytic processing
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spelling Podoplanin is a substrate of presenilin-1/γ-secretaseYurrita, María M.Fernández-Muñoz, BeatrizCastillo, Gaelle delMartín-Villar, EsterRenart, JaimeQuintanilla, MiguelMetalloproteaseγ-SecretasePodoplaninProteolytic processingPodoplanin (PDPN) is a mucin-like transmembrane glycoprotein that plays an important role in development and cancer. Here, we provide evidence that the intracellular domain (ICD) of podoplanin is released into the cytosol following a sequential proteolytic processing by a metalloprotease and γ-secretase. Western blotting and cell fractionation studies revealed that HEK293T and MDCK cells transfected with an eGFP-tagged podoplanin construct (PDPNeGFP, 50-63 kDa) constitutively express two C-terminal fragments (CTFs): a ∼33 kDa membrane-bound PCTF33, and a ∼29 kDa cytosolic podoplanin ICD (PICD). While pharmacological inhibition of metalloproteases reduced the expression of PCTF33, treatment of cells with γ-secretase inhibitors resulted in enhanced PCTF33 levels. PCTF33 processing by γ-secretase depends on presenilin-1 (PS1) function: cells expressing a dominant negative form of PS1 (PS1 D385N), and mouse embryonic fibroblasts (MEFs) genetically deficient in PS1, but not in PS2, show higher levels of PCTF33 expression with respect to wild-type MEFs. Furthermore, transfection of PS1 deficient MEFs with wild-type PS1 (PS1 wt) decreased PCTF33 levels. N-terminal amino acid sequencing of the affinity purified PICD revealed that the γ-secretase cleavage site was located between valines 150 and 151, but these residues are not critical for proteolysis. We found that podoplanin CTFs are also generated in cells expressing podoplanin mutants harboring heterologous transmembrane regions. Taken together, these results indicate that podoplanin is a novel substrate for PS1/γ-secretase. © 2013 Elsevier Ltd. All rights reserved.This work was supported by grants SAF2010-19152 from the Spanish Ministry of Economy and Competitiveness and S2010/BMD-2359 (Skin Model-CM) from the Community of Madrid. M.M.Y. and G.d.C. were funded during part of this work by the Formación de Personal Investigador and Juan de la Cierva programs, respectively. EMV is funded by the Spanish Association for Cancer Research Foundation (AECC).Peer ReviewedElsevierComunidad de MadridMinisterio de Economía y Competitividad (España)Consejo Superior de Investigaciones Científicas (España)Fundación Científica Asociación Española Contra el CáncerConsejo Superior de Investigaciones Científicas [https://ror.org/02gfc7t72]2015201520142015info:eu-repo/semantics/articlehttp://purl.org/coar/resource_type/c_6501Preprintinfo:eu-repo/semantics/submittedVersionhttp://hdl.handle.net/10261/111808reponame:DIGITAL.CSIC. Repositorio Institucional del CSICinstname:Consejo Superior de Investigaciones Científicas (CSIC)Inglés#PLACEHOLDER_PARENT_METADATA_VALUE#S2010/BMD-2359/SKINMODELhttp://dx.doi.org/10.1016/j.biocel.2013.11.016Síinfo:eu-repo/semantics/openAccessoai:digital.csic.es:10261/1118082026-05-22T06:33:51Z
dc.title.none.fl_str_mv Podoplanin is a substrate of presenilin-1/γ-secretase
title Podoplanin is a substrate of presenilin-1/γ-secretase
spellingShingle Podoplanin is a substrate of presenilin-1/γ-secretase
Yurrita, María M.
Metalloprotease
γ-Secretase
Podoplanin
Proteolytic processing
title_short Podoplanin is a substrate of presenilin-1/γ-secretase
title_full Podoplanin is a substrate of presenilin-1/γ-secretase
title_fullStr Podoplanin is a substrate of presenilin-1/γ-secretase
title_full_unstemmed Podoplanin is a substrate of presenilin-1/γ-secretase
title_sort Podoplanin is a substrate of presenilin-1/γ-secretase
dc.creator.none.fl_str_mv Yurrita, María M.
Fernández-Muñoz, Beatriz
Castillo, Gaelle del
Martín-Villar, Ester
Renart, Jaime
Quintanilla, Miguel
author Yurrita, María M.
author_facet Yurrita, María M.
Fernández-Muñoz, Beatriz
Castillo, Gaelle del
Martín-Villar, Ester
Renart, Jaime
Quintanilla, Miguel
author_role author
author2 Fernández-Muñoz, Beatriz
Castillo, Gaelle del
Martín-Villar, Ester
Renart, Jaime
Quintanilla, Miguel
author2_role author
author
author
author
author
dc.contributor.none.fl_str_mv Comunidad de Madrid
Ministerio de Economía y Competitividad (España)
Consejo Superior de Investigaciones Científicas (España)
Fundación Científica Asociación Española Contra el Cáncer
Consejo Superior de Investigaciones Científicas [https://ror.org/02gfc7t72]
dc.subject.none.fl_str_mv Metalloprotease
γ-Secretase
Podoplanin
Proteolytic processing
topic Metalloprotease
γ-Secretase
Podoplanin
Proteolytic processing
description Podoplanin (PDPN) is a mucin-like transmembrane glycoprotein that plays an important role in development and cancer. Here, we provide evidence that the intracellular domain (ICD) of podoplanin is released into the cytosol following a sequential proteolytic processing by a metalloprotease and γ-secretase. Western blotting and cell fractionation studies revealed that HEK293T and MDCK cells transfected with an eGFP-tagged podoplanin construct (PDPNeGFP, 50-63 kDa) constitutively express two C-terminal fragments (CTFs): a ∼33 kDa membrane-bound PCTF33, and a ∼29 kDa cytosolic podoplanin ICD (PICD). While pharmacological inhibition of metalloproteases reduced the expression of PCTF33, treatment of cells with γ-secretase inhibitors resulted in enhanced PCTF33 levels. PCTF33 processing by γ-secretase depends on presenilin-1 (PS1) function: cells expressing a dominant negative form of PS1 (PS1 D385N), and mouse embryonic fibroblasts (MEFs) genetically deficient in PS1, but not in PS2, show higher levels of PCTF33 expression with respect to wild-type MEFs. Furthermore, transfection of PS1 deficient MEFs with wild-type PS1 (PS1 wt) decreased PCTF33 levels. N-terminal amino acid sequencing of the affinity purified PICD revealed that the γ-secretase cleavage site was located between valines 150 and 151, but these residues are not critical for proteolysis. We found that podoplanin CTFs are also generated in cells expressing podoplanin mutants harboring heterologous transmembrane regions. Taken together, these results indicate that podoplanin is a novel substrate for PS1/γ-secretase. © 2013 Elsevier Ltd. All rights reserved.
publishDate 2014
dc.date.none.fl_str_mv 2014
2015
2015
2015
dc.type.none.fl_str_mv info:eu-repo/semantics/article
http://purl.org/coar/resource_type/c_6501
Preprint
info:eu-repo/semantics/submittedVersion
format article
status_str submittedVersion
dc.identifier.none.fl_str_mv http://hdl.handle.net/10261/111808
url http://hdl.handle.net/10261/111808
dc.language.none.fl_str_mv Inglés
language_invalid_str_mv Inglés
dc.relation.none.fl_str_mv #PLACEHOLDER_PARENT_METADATA_VALUE#
S2010/BMD-2359/SKINMODEL
http://dx.doi.org/10.1016/j.biocel.2013.11.016
Sí
dc.rights.none.fl_str_mv info:eu-repo/semantics/openAccess
eu_rights_str_mv openAccess
dc.publisher.none.fl_str_mv Elsevier
publisher.none.fl_str_mv Elsevier
dc.source.none.fl_str_mv reponame:DIGITAL.CSIC. Repositorio Institucional del CSIC
instname:Consejo Superior de Investigaciones Científicas (CSIC)
instname_str Consejo Superior de Investigaciones Científicas (CSIC)
reponame_str DIGITAL.CSIC. Repositorio Institucional del CSIC
collection DIGITAL.CSIC. Repositorio Institucional del CSIC
repository.name.fl_str_mv
repository.mail.fl_str_mv
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score 15,81155

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