Role Of Poly (ADP-ribose) Polymerase 1 In The Regulation Of Gene Transcription In Cardiovascular System

Experimental evidence has demonstrated that reactive oxygen and nitrogen speciesare over-generated in cardiomyocytes,cardiac fibroblasts,smooth muscle cells,endothelial cells,and infiltrating inflammatory cells during a wide range of cardiovasculardiseases,such as circulatory shock,myocardial ischemic-reperfusion injury,myocardialhypertrophy and heart failure,cardiomyopathies,cardiovascular aging,atherosclerosis,and hypertension.Reactive oxygen and nitrogen species induce cellular DNA damage andconsequently activate poly (ADP-ribose) polymerase 1 (PARP-1),a multifunctionalnuclear enzyme present in eukaryotes.Upon binding to DNA damage breaks,PARP-1successively transfer ADP-ribose moiety of nicotinamide dinucleotide (oxidized,NAD~+)to a variety of target proteins.This process of poly(ADP-ribosyl)ation is an importantpost-translational modification of proteins.PARP-1 has been implicated in multiplecellular functions,including DNA repair,maintenance of genomic integrity,cellreplication and differentiation,and transcriptional regulation.A growing body of evidencesuggests that PARP-1 plays an important role in pathogenesis of various cardiovasculardiseases.In related animal models of these diseases,the activation of PARP-1 paticipatesin the transcriptional regulation of numerous genes,which eventually inducecardiovascular dysfunction associated with cardiovascular diseases.Although PARP-1 isimplicated in the transcriptional regulation of many genes through collaboration withtranscription factors,the mechanism underlying PARP-1 regulates transcription of genesinvolved in cardiovascular system,remains unclear.This study focuses on the mechanismthat PARP-1 interacts with several transcription factors AP1,PPARγand NF-κB,toregulate transcription of genes related with cardiovascular system. PartⅠAngiotensinⅡpromotes poly (ADP-ribosyl) ation ofc-Jun/c-Fos in cardiac fibroblastsAims:c-Jun/c-Fos (activator protein 1,AP1) contributes importantly to AngⅡ-inducedcardiac fibrosis through induction of extracellular matrix protein over-expression in cardiacfibroblasts.This study aims to explore the mechanism by which AngⅡpromotesc-Jun/c-Fos transactivation in cultured rat cardiac fibroblasts.Methods and results:In this study,we demonstrated that c-Fos and c-Jun werepoly(ADP-ribosyl)ated in cultured cardiac fibroblasts.Southwestern blot and EMSA assaysshowed that incubation of nuclear extracts with NAD~+ and active DNA increased the basalDNA binding activities of c-Jun (31.0±1.0%,P＜0.01) and AP1 (14.2±3.1%,P＜0.01);incubation of recombinant c-Fos or/and c-Jun with PARP-1,NAD~+ and active DNAincreased the basal DNA binding activities of c-Jun (48.3±4.2%,P＜0.01) and AP1(21.2±1.5%,P＜0.01).Treatment with Ang II promoted PARP-1 activation and enhancedpoly (ADP-ribosyl)ation of c-Fos (14.1+1.1%,P＜0.01) and c-Jun (15.5±5.6%,P＜0.01).AngⅡalso increased the basal DNA binding activities of c-Jun (13.5±2.4%,P＜0.01) andAP1 (18.7±3.5%,P＜0.01) in cultured cells.Inhibition of PARP-1 by PJ34 or siRNAeffectively prevented Ang II-induced increases in the DNA binding of c-Jun and AP1,anddecreased AP1-driven transcription (including collagenⅠα1 andⅢα1,MMP-9 andTIMP-1).Conclusions:This study illustrated that c-Jun and c-Fos were poly(ADP-ribosyl)ated byPARP-1,and poly(ADP-ribosyl)ation enhanced the DNA binding of AP1.Ang II promotedpoly(ADP-ribosyl)ation of c-Jun and c-Fos through activation of PARP-1 and,subsequently,enhanced AP1-driven transcription in cardiac fibroblasts. PartⅡPARP-1 suppresses adiponectin expression throughpoly(ADP-ribosyl)ation of PPARγin cardiac fibroblastsAims:Our aim was to explore the mechanism underlying the transcriptional regulation ofadiponectin and its receptors (AdipoR) in cultured rat cardiac fibroblasts.Methods and results:Using western blot and real-time RT-PCR assays,the expression ofadiponectin and its receptors was determined.Using southwestern blot and electrophoreticmobility shift assays,the DNA binding activity of peroxisome proliferator activatedreceptorγ(PPARγ) was determined.The results showed that adiponectin and AdipoR1were highly expressed in cultured rat cardiac fibroblasts.Inhibition of poly(ADP-ribose)polymerase 1 (PARP-1) by 3-aminobenzamide,PJ34,or PARP-1 siRNA markedlyincreased the transcription of adiponectin and AdipoR1 in cultured fibroblasts,mature 3T3-L1 adipocytes,rat myocardium,and white adipose tissue.PPARγwaspoly(ADP-ribosyl)ated by PARP-1 in cardiac fibroblasts under basal conditions.Poly(ADP-ribosyl)ation of PPARγprevented its binding to DNA.Inhibition of PARP-1enhanced the DNA binding and transactivation of PPARγand increased the transcription ofPPARγ-target genes including CD36,lipoprotein lipase,and leptin in cultured fibroblasts.Conclusions:PARP-1 inhibits adiponectin and AdipoR1 expression as well as PPARγtransactivation through poly (ADP-ribosyl)ation of PPARγin cultured rat cardiacfibroblasts. PartⅢActivation and Overexpression of PARP-1 inCirculating Mononuclear Cells Promote TNF-a and IL-6Expression in Patients with Unstable AnginaAims:Proinflammatory cytokines are involved in the development of unstable angina (UA).Poly(ADP-ribose) polymerase-1 (PARP-1) contributes importantly to regulating the transcriptionof inflammatory cytokines.This study aims to investigate the relationship of PARP-1 in circulatingmononuclear cells (MNCs) and plasma TNF-αand IL-6 in UA patients and to elucidate themechanism that PARP-1 promotes TNF-αand IL-6 expression via NF-κB pathway.Methods and results:Twenty six Braunwald class IIIB UA patients,25 stable anginapatients and 25 healthy volunteers were enrolled in this study.Plasma TNF-αand IL-6were determined with ELISA.Circulating MNCs were analyzed for PARP activity,PARP-1expression and NF-κB DNA binding activity.MNCs from healthy subjects were cultured toinvestigate the direct effects of PARP-1 on NF-κB DNA binding activity and the expressionof TNF-αand IL-6.PARP activity and PARP-1 expression in circulating MNCs wereincreased and positively correlated with plasma TNF-αand IL-6,respectively,in UApatients.Spontaneous NF-κB activation in MNCs was demonstrated in UA patients.Incultured MNCs from healthy subjects,inhibition of PARP-1 preventedlipopolysaccharide-induced increase in DNA binding activity of NF-κB and the expressionof TNF-αand IL-6.Supershift assay demonstrated that PARP-1 was a component ofNF-κB/DNA complex.Addition of recombinant human PARP-1 protein to nuclear extractsof MNCs significantly increased the DNA binding activity of NF-κB.Conclusions:Activation and overexpression of PARP-1 are demonstrated in circulatingMNCs of UA patients.Overexpressed PARP-1 promotes PARP-1/NF-κB/DNA complexformation,thereby enhancing the expression of TNF-αand IL-6 in circulating MNCs ofUA patients.