Wednesday, February 4, 2009

SIRT1


SIRT1 regulates differentiation of mesenchymal stem cells by deacetylating β-catenin.

"[We created] MSC specific SIRT1 knock-out (MSCKO) mice. Aged MSCKO mice (2.2 years old) showed defects in tissues derived from MSCs; i.e. a reduction in subcutaneous fat, cortical bone thickness and trabecular volume. Young mice showed related but less pronounced effects. MSCs isolated from MSCKO mice showed reduced differentiation towards osteoblasts and chondrocytes in vitro, but no difference in proliferation or apoptosis. Expression of β-catenin targets important for differentiation was reduced in MSCKO cells. Moreover, while β-catenin itself (T41A mutant resistant to cytosolic turnover) accumulated in the nuclei of wild-type MSCs, it was unable to do so in MSCKO cells. However, mutating K49R or K345R in β-catenin to mimic deacetylation restored nuclear localization and differentiation potential in MSCKO cells. SIRT1 deacetylates β-catenin to promote its accumulation in the nucleus leading to transcription of genes for MSC differentiation."

"SirT1 heterozygotes show increased osteoarthritis and increased levels of chondrocyte apoptosis in cartilage"

"Cartilage of the long bones in MSCKO mice showed no morphological change"<-so maybe early expression of Beta-Catenin does not inhibit longitudinal bone growth

"when MSCs differentiate, levels of SIRT1 decrease, and acetylation of β-catenin increases"

I couldn't access the supplementary data files and it looked like there was some good info in there.

Set7/9 impacts COL2A1 expression through binding and repression of SirT1 histone deacetylation.

"COL2A1 gene expression is positively regulated by the NAD-dependent protein deacetylase SirT1, through its ability to bind chromatin regions of the COL2A1 promoter and enhancer. Human chondrocytes were encapsulated in three-dimensional (3D) alginate beads where they exhibited up-regulated COL2A1 mRNA expression and increased levels of SirT1 occupancy on the promoter and enhancer regions, when compared to monolayer controls. Chromatin immunoprecipitation (ChIP) analyses of 3D cultures showed augmentated levels of the DNA-binding transcription factor SP1, and the histone methyltransferase Set7/9, on the COL2A1 promoter site. ChIP reChIP assays revealed that SirT1 and Set7/9 form a protein complex on the COL2A1 promoter region of 3D-cultured chondrocytes, which also demonstrated elevated trimethylated lysine 4 on histone 3 (3MeH3K4), a hallmark of Set7/9 methyltransferase activity. Advanced passaging of chondrocytes yielded a decrease in 3MeH3K4 and Set7/9 levels on the COL2A1 promoter and reduced COL2A1 expression, suggesting that the SirT1/Set7/9 complex is preferentially formed on the COL2A1 promoter and required for gene activation. Interestingly, despite SirT1 occupancy, its deacetylation targets (i.e H3K9/14 and H4K16) were found acetylated on the COL2A1 promoter of 3D-cultured chondrocytes. A possible explanation for this phenotype is the enrichment of the histone acetyltransferases P300 and GCN5 on the COL2A1 promoter of3 D-cultured chondrocytes. Our study indicates that Set7/9 prevents the histone deacetylase activity of SirT1, potentiating euchromatin formation on the promoter site of COL2A1 and resulting in morphology-dependent COL2A1 gene transactivation."

"human chondrocyte cell lines stably overexpressing SirT1 displayed increased expression of COL2A1 mRNA and demonstrate enriched occupancy of SirT1, Sox9 and PGC1α on the gene enhancer region. Similarly, TNFα-stimulated human chondrocytes, which possessed reduced COL2A1 mRNA expression, displayed an inactive SirT1 cleaved variant (75SirT1) that yielded reduced occupancy of PGC1α and Sox9 on the COL2A1 enhancer region"

"chondrocyte passaging reduces cellular SirT1 activity, which is likely a result of increased inhibitory DBC1."

"SirT1 causes Set7/9 association on the COL2A1 promoter, leading to increased levels of 3MeH3K4."

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