Abstract 3605: BAT1 is an Important Regulator of Protein Synthesis in Cardiomyocytes
BAT1, an ATP dependent RNA helicase that also has ATPase activity, is a member of the DExD/H box family and is phylogenetically grouped with the elongation initiation factor eIF4A, the prototypical member of the DExD/H box family of helicases. BAT1, also known as UAP56, is an essential RNA splicing factor required for spliceosome assembly and mRNA export but its role in protein synthesis is unknown. Binding of ATP to lysine 95 (K95) of BAT1 is required for mRNA export from the nucleus to the cytoplasm. Mutation of K95 inhibits BAT1’s ATPase and helicase activity. Aspartic acid 199 (D199) is critical for BAT1’s helicase activity and mutation of D199 is associated with inhibition of BAT1’s helicase activity but not its ATPase activity. Given the role of BAT1 in RNA splicing and mRNA export, we hypothesized that BAT1 plays an important role in protein synthesis in cardiomyocytes and subsequent hypertrophy. Wild type (WT) BAT1, K95M BAT1 mutant, or D199A BAT1 mutant were over-expressed in HeLa cells and protein synthesis was measured by leucine incorporation assay. We found that WT BAT1 increased protein synthesis compared with control (2.0±0.27 fold, p<0.01). BAT1 K95M mutant and D199A mutant both inhibited protein synthesis compared with WT BAT1 (relative leucine incorporation 1.15±0.40 vs. 3.15±0.55 and 2.13±0.44 vs. 3.15±0.55, respectively, p<0.01) suggesting that the RNA helicase activity of BAT1 is critical for protein synthesis. Because protein synthesis is a key component of hypertrophy, we examined the role of BAT1 in regulating protein synthesis in cardiomyocytes. We found that BAT1 siRNA inhibited phenylephrine (PE) induced protein synthesis in cardiomyocytes (62.6±5.5% inhibition, p<0.01). Immunofluorescence staining demonstrated that BAT1 is localized inside the nucleus. There was no increase in BAT1 expression in cardiomyocytes following treatment with Angiotensin II or with PE. Our preliminary data suggest that BAT1 siRNA inhibits PE induced cardiomyocyte hypertrophy. NF-κB is a regulator of myocyte growth but we found that BAT1 had no effect on NF-κB activity in cardiomyocytes. In conclusion, our data demonstrate that BAT1 is an important regulator of protein synthesis and may have an important role in the regulation of cardiac hypertrophy.