Abstract 352: DnaJb5, an Hsp40 Family Protein, Mediates Antihypertrophic Effects of Thioredoxin1 in the Heart
Thioredoxin1 (Trx1) reduces various redox-sensitive proteins and regulates cell growth and death. Cardiac hypertrophy induced by pressure-overload is suppressed in transgenic mice with cardiac specific overexpression of Trx1 (Tg-Trx1). To identify the molecular mechanism by which Trx1 mediates antihypertrophic effects, we employed cDNA microarray analyses. The result indicated that DnaJb5, a heat shock protein 40 (Hsp40) family protein, is significantly upregulated in Tg-Trx1 hearts. Protein expression of DnaJb5 was increased in both Trx1-overexpressing myocytes and Tg-Trx1 (184% and 372%, p<0.05, respectively). The upregulation is mediated by Heat shock factor-1 (HSF-1), since both expression and transcriptional activity of HSF-1 were enhanced in Trx1-overexpressed myocytes by 280% and 187% (p<0.05), respectively. Furthermore, shRNA-mediated knockdown of HSF-1 attenuated Trx1-induced upregulation of DnaJb5 (51%, p<0.05). Immunostaining and western blot analyses indicated that Trx1 and DnaJb5 are colocalized in the nucleus of myocytes. Pull-down assays indicated that DnaJb5 associates with Trx1 through TXNIP, a Trx1-binding protein. Knockdown of Trx1 reduced the amount of Trx1 and DnaJb5 in the nucleus. Pull-down assays also showed that DnaJb5 interacts directly with histone deacetylase 4 (HDAC4). Whereas overexpression of Trx1 induced HDAC4 accumulation in the nucleus, knockdown of Trx1 by shRNA excluded HDAC4 from the nucleus. Trx1 and DnaJb5 attenuated phenylephrine (PE)-induced increases in the transcriptional activity of NFAT (54% and 59%, p < 0.05, respectively) and myocyte hypertrophy without showing additive effects. Furthermore, knockdown of DnaJb5 significantly attenuated the Trx1-mediated inhibition of PE-induced hypertrophy. In a genetic cross between Tg-Trx1 and transgenic mice harboring the NFAT luciferase reporter, both NFAT activation and cardiac hypertrophy induced by PE were significantly suppressed in the presence of Trx1. Collectively, Trx1 upregulates DnaJb5 via HSF-1, colocalizes with DnaJb5 in the nucleus, and inhibits NFAT activity through recruitment of HDAC4 into the nucleus. DnaJb5 is a critical downstream effector of Trx1, mediating anti-hypertrophic effects of Trx1 in the heart.