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dc.contributor.authorGalindo, Antonio-
dc.contributor.authorCalcagno-Pizarelli, Ana María-
dc.contributor.authorArst, Herbert Nathan Jr.-
dc.contributor.authorPeñalva, Miguel Ángel-
dc.identifier.citationJournal of Cell Science 125:1784-1795(2012)es_ES
dc.description12 páginas, 9 figuras -- PAGS nros. 1784-1795es_ES
dc.description.abstractThe fungal pal/RIM signalling pathway, which regulates gene expression in response to environmental pH involves, in addition to dedicated proteins, several components of ESCRT complexes, which suggested that pH signalling proteins assemble on endosomal platforms. In Aspergillus nidulans, dedicated Pal proteins include the plasma membrane receptor PalH and its coupled arrestin, PalF, which becomes ubiquitylated in alkaline pH conditions, and three potentially endosomal ESCRT-III associates, including Vps32 interactors PalA and PalC and Vps24 interactor calpain-like PalB. We studied the subcellular locations at which signalling takes place after activating the pathway by shifting ambient pH to alkalinity. Rather than localising to endosomes, Vps32 interactors PalA and PalC transiently colocalise at alkaline-pH-induced cortical structures in a PalH-, Vps23- and Vps32-dependent but Vps27-independent manner. These cortical structures are much more stable when Vps4 is deficient, indicating that their half-life depends on ESCRT-III disassembly. Pull-down studies revealed that Vps23 interacts strongly with PalF, but co-immunoprecipitates exclusively with ubiquitylated PalF forms from extracts. We demonstrate that Vps23–GFP, expressed at physiological levels, is also recruited to cortical structures, very conspicuous in vps27Δ cells in which the prominent signal of Vps23–GFP on endosomes is eliminated, in a PalF- and alkaline pH-dependent manner. Dual-channel epifluorescence microscopy showed that PalC arrives at cortical complexes before PalA. As PalC recruitment is PalA independent and PalA recruitment is PalC dependent but PalB independent, these data complete the participation order of Pal proteins in the pathway and strongly support a model in which pH signalling takes place in ESCRT-containing, plasma-membrane-associated, rather than endosome-associated, complexeses_ES
dc.description.sponsorshipThis work was supported by Ministerio de Ciencia [grant number BIO2009-7281 to M.A.P.]; Comunidad de Madrid [grant number SAL/0246/2006 to M.A.P.]; the Biotechnology and Biological Sciences Research Council [grant numbers BB/D521781/1, BB/F01189X/1 to H.N.A. and, for the latter, Elaine Bignell]; and the Wellcome Trust [grant numbers 067878, 084660/Z/08/Z to H.N.A. and Joan Tilburnes_ES
dc.publisherCompany of Biologistses_ES
dc.subjectSeven transmembrane receptores_ES
dc.subjectMultivesicular body pathwayes_ES
dc.subjectSignal transductiones_ES
dc.subjectAspergillus nidulanses_ES
dc.titleAn ordered pathway for the assembly of ESCRT-containing fungal ambient pH signalling complexes at the plasma membranees_ES
dc.description.peerreviewedPeer reviewedes_ES
dc.relation.publisherversionhttp://dx.doi.org/ 10.1242/​jcs.098897es_ES
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