1.SIRT1Links CIITA Deacetylation To MHC Ⅱ Activation

Objectives: Antigen-dependent stimulation of T cells plays a critical role in adaptiveimmunity and host defense.Activation of major histocompatibility complex II (MHCII) molecules, dictated by Class II transactivator (CIITA), is considered a pivotal stepin this process. The mechanism underlying differential regulation of CIITA activity bythe post-translational modification machinery (PTM) and its implications are notclearly appreciated.As a NAD+-dependent deacetylase, SIRT1, by deacetylating histones and moreoften non-histone protein factors, has been implicated in a number of physiologicaland pathological processes.We have previously demonstrated that HDAC2, a class Ideacetylase, interacts with CIITA and suppresses its activity by targeting it toproteasomal degradation. Several recent investigations have implicated the class IIIdeacetylase SIRT1in regulating the immune system, therefore we set to determinewhether MHC II transcription, a key pathophysiological process controlled by CIITA,was affected by SIRT1.Methods: when SIRT1is overexpressed or silenced, mRNA levels of MHC II wereexamined by real-time quantitative PCR (qPCR) in human embryonic kidney cell(293), human leukemia monocytic/macrophage (THP-1). Luciferase reporter assayswere performed to assess the effect of SIRT1on the promoter activities ofCIITA.Immunoprecipitation and western blotting were performed to detect theassociation between SIRT1and CIITA,as well as the acetylation level of CIITA.Theeffect of SIRT1on prolonging CIITA half-life were detected by using Westernblotting.ChIP assay were performed to examine the effect of SIRT1on promotingCIITA binding to the promoter of MHC II.When NAMPT was overexpressed,thetranscriptional level of MHC II was examined by luciferase reporter assay and theacetylation level of CIITA was detected by using IP.Finally, under hypoxia and ox-LDL stress,we examined the acetylation level of CIITA by usingimmunoprecipitation.ChIP assay and luciferase reporter assays were performed toassess MHC II transcriptional regulation by CIITA.Results: SIRT1potentiats the activation of MHC II transcription by CIITA inmacrophages.The underlying mechanism:SIRT1can interact with and deacetylatesCIITA,thus will augment the stability and promoter binding of CIITA.SIRT1activity depends on intracellular levels of NAD+,which is synthesizedprimarily by NAM phosphoribosyltransferase (NAMPT).NAMPT contributes toCIITA-dependent MHC II transactivation by upregulating SIRT1activity.Whereas thestress of hypoxia and oxLDL suppresses CIITA-dependent transcriptional activationof MHC IIgenes via attenuating the expression and activity of SIRT1in macrophages.Conclusion: As a NAD+-dependent deacetylase,SIRT1interact with and deacetylatesCIITA.,thus will augment the stability and promoter binding of CIITA. Thus, our datalink SIRT1-mediated deacetylation of CIITA to MHC II transactivation inmacrophages and highlight a novel strategy stress cues may employ to manipulatehost adaptive immune system. Objectives: Cardiovascular disease (CVD) is the leading cause of death worldwide.Epidemiological studies have pointed to a persistent increase in CVD-releatedmorbidity and mortality in the past decade with the alteration of life style and dietarychoices. Myocardial infarction (MI), which consistutes a major component of CVD,results from the interruption of blood supply to the heart causing necrosis ofcardiomyocyte. This is most commonly due to occlusion of a coronary arteryfollowing the rupture of a vulnerable atherosclerotic plaque. With the advancement ofmedical sciences, considerable progress has been made in the fundamental research ofCVD represented by a shift from invesigations that focus on anatomy and wholisticphysiology to investigations that center on cell and molecular biology.It has been documented that SIRT1protects mice against MI and that SIRT1expression is down-regulated by hypoxic ischemic stress although the mechanismunderlying repression of SIRT1transcription is not clearly understood. We examinedthe role of the histone H3K9trimethylase SUV39h2in the transcriptional repressionof SIRT1during MI.When human embryonic kidney cells (293) and rat cardiac myoblast (H9C2) weretreated with MTA (a pan-methyltransferase inhibitor), mRNA level of SIRT1wasexamined by real-time quantitative PCR (qPCR) while its transcriptional activity wasdetected by luciferase reporter assays. In addition, Western blotting was performed toexamine the protein expression of SIRT1. Silencing of SUV39h expression wasmediated by siRNA. Inhibition of SUV39h activity was achieved by treatment with aspecific inhibitor, chaetocin. The effect of chaetocin on MI was assessed in vivo in amouse model by permanent ligation of the left anterior descending coronary.Results: In H9C2cells, adminstration of MTA and chaetocin augmented SIRT1transcription and expression. More importantly, chaetocin treatment alleviated therepression of SIRT1expression under hypoxic ischemic stress. Depletion of SUV39h2, but not SUV39h1, exerted similar effects on SIRT1expression aschaetocin. Finally, pre-treatment with chaetocin significantly attenuated MI in mice.Therefore, targeting SUV39h2by chaetocin may yield a viable strategy in therapeuticintervention of MI.