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Conference Paper: Sirtuin 1 interacts with DNA repair molecules during the initiation phase of reprogramming from mouse fibroblasts to mouse induced pluripotent stem cells

TitleSirtuin 1 interacts with DNA repair molecules during the initiation phase of reprogramming from mouse fibroblasts to mouse induced pluripotent stem cells
Authors
Issue Date2014
PublisherThe International Society for Stem Cell Research (ISSCR).
Citation
The 12th Annual Meeting of the International Society for Stem Cell Research (ISSCR), Vancouver, Canada, 28-21 June 2014. In the Abstracts of the 12th Annual Meeting of the International Society for Stem Cell Research (ISSCR), 2014, p. 446, abstract no. F-2192 How to Cite?
AbstractThe reprogramming of somatic cells to induced pluripotent stem cells (iPSCs) involves chromatin remodeling. We have previously shown that a NAD+ dependent histone deacetylase, sirtuin 1 (SIRT1) positively regulated mouse iPSCs formation, partly through deacetylating p53 leading to altered Nanog and p21 expression (1). SIRT1 also deacetylates other proteins like PGC1α [2] and FOXO3 [3]. The present study aimed at identifying the SIRT1 interacting protein during the reprogramming process. Pure population of secondary mouse embryonic fibroblast (MEF) containing doxycycline (DOX) inducible OSKM factors were obtained from chimeric MEF by Fluorescence Activated Cell Sorting. The cells were cultured in the presence of DOX for the first 5 days (initiation phase of reprogramming). The interacting proteins of SIRT1 during reprogramming were isolated by co-immunoprecipitation. The immunoprecipitated proteins were visualized by silver staining, and identified by mass spectrometry. According to the pathway clustering analysis from the Kyoto Encyclopedia of Genes and Genomes (KEGG), a number of them were involved in repair of double strand break (DSB). Homologous recombination DNA repair genes are important for successful reprogramming [4]. We speculated that SIRT1 might promote DNA repair through binding to the complex of DSB repair proteins. Indeed, the levels of the SIRT1 interacting DSB repair proteins (SIRT1-DSB) were up-regulated in the MEF upon DOX induction. To this end, we tried to knock down two of these SIRT1-DSBs at the onset of reprogramming. The result showed that the down-regulation of SIRT1-DSBs suppressed the iPSC colony formation as demonstrated by lower colony number and reduced Nanog expressions in the colonies. These data supported that SIRT1 activated the SIRT1-DSBs in the initial phase of reprogramming process leading to increase in colony formation. Acknowledgement: This work is supported by General Research Fund (HKU775711M) and Seed Funding Scheme to Support Research Projects on Human Embryonic Stem Cells (ESC) and Induced Pluripotent Stem Cells (iPSC), Stem Cell and Regenerative Medicine Consortium (SCRMC), Li Ka Shing Faculty of Medicine, The University of Hong Kong. We thank Professor Andras Nagy (The University of Toronto) for his generous gift of primary iPSCs.
DescriptionPoster Session: iPS Cells
Persistent Identifierhttp://hdl.handle.net/10722/204889

 

DC FieldValueLanguage
dc.contributor.authorLee, CYLen_US
dc.contributor.authorPeng, Qen_US
dc.contributor.authorChen, CHen_US
dc.contributor.authorNg, EHYen_US
dc.contributor.authorYeung, WSBen_US
dc.date.accessioned2014-09-20T00:56:01Z-
dc.date.available2014-09-20T00:56:01Z-
dc.date.issued2014en_US
dc.identifier.citationThe 12th Annual Meeting of the International Society for Stem Cell Research (ISSCR), Vancouver, Canada, 28-21 June 2014. In the Abstracts of the 12th Annual Meeting of the International Society for Stem Cell Research (ISSCR), 2014, p. 446, abstract no. F-2192en_US
dc.identifier.urihttp://hdl.handle.net/10722/204889-
dc.descriptionPoster Session: iPS Cells-
dc.description.abstractThe reprogramming of somatic cells to induced pluripotent stem cells (iPSCs) involves chromatin remodeling. We have previously shown that a NAD+ dependent histone deacetylase, sirtuin 1 (SIRT1) positively regulated mouse iPSCs formation, partly through deacetylating p53 leading to altered Nanog and p21 expression (1). SIRT1 also deacetylates other proteins like PGC1α [2] and FOXO3 [3]. The present study aimed at identifying the SIRT1 interacting protein during the reprogramming process. Pure population of secondary mouse embryonic fibroblast (MEF) containing doxycycline (DOX) inducible OSKM factors were obtained from chimeric MEF by Fluorescence Activated Cell Sorting. The cells were cultured in the presence of DOX for the first 5 days (initiation phase of reprogramming). The interacting proteins of SIRT1 during reprogramming were isolated by co-immunoprecipitation. The immunoprecipitated proteins were visualized by silver staining, and identified by mass spectrometry. According to the pathway clustering analysis from the Kyoto Encyclopedia of Genes and Genomes (KEGG), a number of them were involved in repair of double strand break (DSB). Homologous recombination DNA repair genes are important for successful reprogramming [4]. We speculated that SIRT1 might promote DNA repair through binding to the complex of DSB repair proteins. Indeed, the levels of the SIRT1 interacting DSB repair proteins (SIRT1-DSB) were up-regulated in the MEF upon DOX induction. To this end, we tried to knock down two of these SIRT1-DSBs at the onset of reprogramming. The result showed that the down-regulation of SIRT1-DSBs suppressed the iPSC colony formation as demonstrated by lower colony number and reduced Nanog expressions in the colonies. These data supported that SIRT1 activated the SIRT1-DSBs in the initial phase of reprogramming process leading to increase in colony formation. Acknowledgement: This work is supported by General Research Fund (HKU775711M) and Seed Funding Scheme to Support Research Projects on Human Embryonic Stem Cells (ESC) and Induced Pluripotent Stem Cells (iPSC), Stem Cell and Regenerative Medicine Consortium (SCRMC), Li Ka Shing Faculty of Medicine, The University of Hong Kong. We thank Professor Andras Nagy (The University of Toronto) for his generous gift of primary iPSCs.en_US
dc.languageengen_US
dc.publisherThe International Society for Stem Cell Research (ISSCR).-
dc.relation.ispartofAnnual Meeting of the International Society for Stem Cell Research (ISSCR)en_US
dc.titleSirtuin 1 interacts with DNA repair molecules during the initiation phase of reprogramming from mouse fibroblasts to mouse induced pluripotent stem cellsen_US
dc.typeConference_Paperen_US
dc.identifier.emailLee, CYL: cherielee@hku.hken_US
dc.identifier.emailNg, EHY: nghye@hku.hken_US
dc.identifier.emailYeung, WSB: wsbyeung@hkucc.hku.hken_US
dc.identifier.authorityLee, CYL=rp00308en_US
dc.identifier.authorityNg, EHY=rp00426en_US
dc.identifier.hkuros237526en_US
dc.identifier.spage446, abstract no. F-2192-
dc.identifier.epage446, abstract no. F-2192-
dc.publisher.placeUnited States-

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