Anti-ATP5B monoclonal antibody, clone 5.4F9.E21

Intramuscular fat (IMF) content is a key determinant of beef quality, making it a key topic of research interest. ATP5B serves as the catalytic component of the mitochondrial ATP synthase complex and plays essential roles in controlling fat contents and oxidative metabolism in bovine skeletal muscle. In this study, we determined that bovine ATP5B was highly expressed in longissimus thoracis. To elucidate the molecular mechanisms involved in bovine ATP5B regulation, we cloned and characterized the promoter region of ATP5B. Applying 5'-rapid amplification of cDNA end analysis (RACE), we identified two transcriptional start sites (TSSs) in its promoter region. Using a series of 5'-deletion promoter plasmids in luciferase reporter assay, we found that the proximal minimal promoter of ATP5B was located within the region -539/220 relative to the TSS. Site-directed mutation in combination with chromatin immunoprecipitation (ChIP) assays demonstrated that MyoD and GATA1 binding to the promoter region drives bovine ATP5B transcription. Taken together, these results provide new insight into the regulatory mechanisms of ATP5B transcription in mediating the IMF content of beef.

Although gene expression following bortezomib treatment has been previously explored, direct effects of bortezomib-induced proteasome inhibition on protein level has not been analyzed so far. Using 2-D PAGE in five mantle cell lymphoma cell lines, we screened for cellular protein level alterations following treatment with 25 nM bortezomib for up to 4 h. Using MS, we identified 38 of the 41 most prominent reliably detected protein spots. Twenty-one were affected in all cell lines, whereas the remaining 20 protein spots were exclusively altered in sensitive cell lines. Western blot analysis was performed for 17 of the 38 identified proteins and 70.6% of the observed protein level alterations in 2-D gels was verified. All cell lines exhibited alterations of the cellular protein levels of heat shock-induced protein species (HSPA9, HSP7C, HSPA5, HSPD1), whereas sensitive cell lines also displayed altered cellular protein levels of energy metabolism (ATP5B, AK5, TPI1, ENO-1, ALDOC, GAPDH), RNA and transcriptional regulation (HNRPL, SFRS12) and cell division (NEBL, ACTB, SMC1A, C20orf23) as well as tumor suppressor genes (ENO-1, FH). These proteins clustered in a tight interaction network centered on the major cellular checkpoints TP53. The results were confirmed in primary mantle cell lymphoma, thus confirming the critical role of these candidate proteins of proteasome inhibition.