Exploring Novel Functions Of SIRT1 In Retinoic Acid Signaling And Stem Cell Bioloby

Retinoid homeostasis is critical for embryonic development, both the deficiency and excess of Retinoic acid(RA) are associated with congenital malformations and developmental defects. SIRT1, the most conserved mammalian NAD+-dependent deacetylase, is an important cellular metabolic sensor crucially involved in multiple cellular processes, extending mammalian lifespan and affecting diseases related to metabolism and aging. Knocking out SIRT1 systemically in mice leads to severe developmental defects in multiple tissues. However, the molecular mechanisms underlying the important developmental phenotype of SIRT1 knockout mice are very unclear.Here we show that SIRT1 is critically involved in the maintenance of homeostatic RA signaling and modulates mouse embryonic stem cell(m ESC) differentiation in part through deacetylation of cellular retinoic acid binding protein Ⅱ(CRABPⅡ), and that hyperactived RA signaling contributes to the developmental defects of SIRT1 KO mice. Using a Stable Isotope Labeling by Amino Acids in Cell Culture(SILAC)-based lys-acetylomic method, we identify CRABPⅡ, a key mediator of RA signaling, as a hyper-acetylated protein in SIRT1 deficient MEFs. Our results show that CRABPⅡ is a novel deacetylation substrate of SIRT1. SIRT1 interacts with and deacetylates CRABPⅡ at K102 site after the RA treatment, reducing its nuclear localization and inhibiting subsequent RAR activation both in cells and in vivo. Consequently, SIRT1 deficiency enhances RA signaling and leads to significantly increased differentiation morphology and dramatically increased differentiation markers upon RA treatment in the m ESCs. Microarray data further support the notion that SIRT1 is crucial for maintenance of ES cell pluripotency. Moreover, silencing or deacetylating CRABPⅡ could partly rescue the phenotype caused by SIRT1 deficiency, indicating that SIRT1 deficiency accelerates RA-induced m ESC differentiation in part through CRABPⅡ. Finally, multiple SIRT1 deficiency induced developmental defects, including delayed mineralization process of endochondral ossification, are associated with elevated RA signaling in mice.Our findings reveal a novel molecular mechanism that regulates cellular RA signaling, and highlight the importance of SIRT1 in transcriptional regulation of ESC pluripotency and embryogenesis. Since SIRT1 is a key metabolic sensor which is hypersensitive to the cellular metabolic and environmental signals, this newly characterized SIRT1/CRABPⅡ/RAR signaling cascade will provide a new avenue to study gene-environment interactions that affect animal development.