Lu Jing, Zhang Renwei, Hong Huiqi, Yang Zuolong, Sun Duanping, Sun Shuya, Guo Xiaolei, Ye Jiantao, Li Zhuoming, Liu Peiqing
Department of Pharmacology and Toxicology, School of Pharmaceutical Sciences, Sun Yat-Sen University, Guangzhou 510006, Guangdong, PR China; National and Local United Engineering Lab of Druggability and New Drugs Evaluation, School of Pharmaceutical Sciences, Sun Yat-Sen University, Guangzhou 510006, Guangdong, PR China; Guangdong Provincial Key Laboratory of Construction Foundation, School of Pharmaceutical Sciences, Sun Yat-Sen University, Guangzhou 510006, Guangdong, PR China.
National and Local United Engineering Lab of Druggability and New Drugs Evaluation, School of Pharmaceutical Sciences, Sun Yat-Sen University, Guangzhou 510006, Guangdong, PR China.
Biochim Biophys Acta. 2016 Dec;1863(12):3027-3039. doi: 10.1016/j.bbamcr.2016.09.019.
The Forkhead box-containing protein, O subfamily 3 (FoxO3) transcription factor negatively regulates myocardial hypertrophy, and its transcriptional activity is finely conditioned by diverse posttranslational modifications, such as phosphorylation, acetylation, ubiquitination, methylation and glycosylation. Here, we introduce a novel modification of the FoxO3 protein in cardiomyocytes: poly(ADP-ribosyl)ation (PARylation) mediated by poly(ADP-ribose) polymerase-1 (PARP1). This process catalyzes the NAD-dependent synthesis of polymers of ADP-ribose (PAR) and their subsequent attachment to target proteins by PARPs. Primary-cultured neonatal rat cardiomyocytes were incubated with isoproterenol (ISO) to induce hypertrophy, or were infected with recombinant adenovirus vectors harboring PARP1 cDNA (Ad-PARP1). Sprague-Dawley (SD) rats were treated with ISO to induce cardiac hypertrophy, or were injected with Ad-PARP1 into the anterior and posterior left ventricular walls. Cardiomyocyte surface area, the mRNA expression of hypertrophic biomarkers, echocardiography, morphometry of the hearts were measured. The PARP1 activity was tested by cellular PAR levels. Interactions of PARP1 and FoxO3 were investigated by co-immunoprecipitation and immunofluorescence technique. PARylation of FoxO3 mediated by PARP1 facilitated its phosphorylation at the T32, S252 and S314 sites, triggered its nucleus export and suppressed its transcriptional activity and target genes expression, ultimately inducing cardiac hypertrophy. Additionally, PARP1 silencing or specific inhibition by 3-Aminobenzamide (3AB) and veliparib (ABT-888) alleviated the inhibition of FoxO3 activity by ISO, thus suppressing ISO-induced cardiac hypertrophy. Our data provide the first evidence that PARP1 exacerbates cardiac hypertrophy by PARylation of FoxO3.
含叉头框蛋白O亚家族3（FoxO3）转录因子对心肌肥大起负调控作用，其转录活性受到多种翻译后修饰的精细调节，如磷酸化、乙酰化、泛素化、甲基化和糖基化。在此，我们介绍心肌细胞中FoxO3蛋白的一种新型修饰：由聚（ADP-核糖）聚合酶-1（PARP1）介导的聚（ADP-核糖）化（PARylation）。该过程催化依赖NAD的ADP-核糖（PAR）聚合物的合成，并随后由PARP将其连接到靶蛋白上。将原代培养的新生大鼠心肌细胞用异丙肾上腺素（ISO）孵育以诱导肥大，或用携带PARP1 cDNA的重组腺病毒载体（Ad-PARP1）感染。用ISO处理Sprague-Dawley（SD）大鼠以诱导心脏肥大，或将Ad-PARP1注射到左心室前壁和后壁。测量心肌细胞表面积、肥大生物标志物的mRNA表达、超声心动图、心脏形态学。通过细胞PAR水平检测PARP1活性。通过免疫共沉淀和免疫荧光技术研究PARP1与FoxO3的相互作用。PARP1介导的FoxO3的PARylation促进其在T32、S252和S314位点的磷酸化，触发其核输出并抑制其转录活性和靶基因表达，最终诱导心脏肥大。此外，PARP1沉默或用3-氨基苯甲酰胺（3AB）和维利帕尼（ABT-888）进行特异性抑制可减轻ISO对FoxO3活性的抑制，从而抑制ISO诱导的心脏肥大。我们的数据首次证明PARP1通过FoxO3的PARylation加剧心脏肥大。
