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TriPer, an optical probe tuned to the endoplasmic reticulum tracks changes in luminal H2O2

2012-06Journal article Open access
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dc.contributor.authorMelo, Eduardo
dc.contributor.authorRafael dos Reis Lopes, Carlos
dc.contributor.authorGollwitzer, Peter
dc.contributor.authorLortz, Stephan
dc.contributor.authorLenzen, Sigurd
dc.contributor.authorMehmeti, Ilir
dc.contributor.authorKaminski, Clemens F.
dc.contributor.authorRon, David
dc.contributor.authorAvezov, Edward
dc.date.accessioned2018-12-07T14:53:29Z
dc.date.available2018-12-07T14:53:29Z
dc.date.issued2012-06
dc.description.abstractBackground: The fate of hydrogen peroxide (H2O2) in the endoplasmic reticulum (ER) has been inferred indirectly from the activity of ER-localized thiol oxidases and peroxiredoxins, in vitro, and the consequences of their genetic manipulation, in vivo. Over the years hints have suggested that glutathione, puzzlingly abundant in the ER lumen, might have a role in reducing the heavy burden of H2O2 produced by the luminal enzymatic machinery for disulfide bond formation. However, limitations in existing organelle-targeted H2O2 probes have rendered them inert in the thiol-oxidizing ER, precluding experimental follow-up of glutathione's role in ER H2O2 metabolism. Results: Here we report on the development of TriPer, a vital optical probe sensitive to changes in the concentration of H2O2 in the thiol-oxidizing environment of the ER. Consistent with the hypothesized contribution of oxidative protein folding to H2O2 production, ER-localized TriPer detected an increase in the luminal H2O2 signal upon induction of pro-insulin (a disulfide-bonded protein of pancreatic beta-cells), which was attenuated by the ectopic expression of catalase in the ER lumen. Interfering with glutathione production in the cytosol by buthionine sulfoximine (BSO) or enhancing its localized destruction by expression of the glutathione-degrading enzyme ChaC1 in the lumen of the ER further enhanced the luminal H2O2 signal and eroded beta-cell viability. Conclusions: A tri-cysteine system with a single peroxidatic thiol enables H2O2 detection in oxidizing milieux such as that of the ER. Tracking ER H2O2 in live pancreatic beta-cells points to a role for glutathione in H2O2 turnover.
dc.description.sponsorshipWellcome Trust [Wellcome 200848/Z/16/Z, WT: UNS18966]; Fundacao para a Ciencia e Tecnologia, Portugal [PTDC/QUI/BIQ/119677/2010, UID/BIM/04773/2013-CBMR]; European Commission (FU FP7 Beta-Bat) [277713]; EPSRC [1503478]; MRC [MR/K015850/1]; Wellcome Trust Strategic Award for core facilities to the Cambridge Institute for Medical Research [Wellcome 100140]; Wellcome Trust Principal Research Fellow
dc.identifier.doi10.1186/s12915-017-0367-5
dc.identifier.issn1741-7007
dc.identifier.urihttp://hdl.handle.net/10400.1/11538
dc.language.isoeng
dc.peerreviewedyes
dc.publisherBiomed Central Ltd
dc.subjectDisulfide-Bond Formation
dc.subjectEnzyme Gene-Expression
dc.subjectHydrogen-Peroxide
dc.subjectThiol-Redox
dc.subjectBeta-Cell
dc.subjectGlutathione
dc.subjectProtein
dc.subjectPeroxiredoxin
dc.subjectStress
dc.subjectPoise
dc.titleTriPer, an optical probe tuned to the endoplasmic reticulum tracks changes in luminal H2O2
dc.typejournal article
dspace.entity.typePublication
oaire.citation.titleBmc Biology
oaire.citation.volume15
person.familyNamePinho Melo
person.familyNameLopes
person.givenNameEduardo
person.givenNameCarlos
person.identifier1443188
person.identifier.ciencia-id3C1C-C10C-1510
person.identifier.ciencia-id5B12-908A-21A2
person.identifier.orcid0000-0002-0974-8977
person.identifier.orcid0000-0002-6136-0478
person.identifier.ridK-4979-2015
person.identifier.scopus-author-id35566177900
rcaap.rightsopenAccess
rcaap.typearticle
relation.isAuthorOfPublication5fa1895f-5577-4652-961a-886ec9bf41b1
relation.isAuthorOfPublication21cb74bb-fd65-4da8-a95c-73d711c9c9d3
relation.isAuthorOfPublication.latestForDiscovery5fa1895f-5577-4652-961a-886ec9bf41b1

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