The current study examined angiotensin receptor (ATR) regulation in proliferating rat aortic vascular smooth muscle cells (VSMCs) in culture. Radioligand competition analysis coupled with RNase protection assays (RPAs) revealed that angiotensin type 1a receptor (AT1aR) densities (Bmax) increased by 30% between 5 and 7 days in culture [Bmax (fmol/mg protein): day 5, 379 ± 8.4 vs. day 7, 481 ± 12, n = 3, P < 0.05] under conditions in which no significant changes in AT1aR mRNA expression occurred [in RPA arbitrary units (AU): day 5, 0.23 ± 0.01 vs. day 7, 0.24 ± 0.04, n = 4] or in mRNA synthesis determined by nuclear run-on assays [AU: day 5, 0.35 ± 0.14 vs. day 7, 0.33 ± 0.11, n = 5]. In contrast, polysome distribution analysis indicated that AT1aR mRNA was more efficiently translated in day 7 cells compared with day 5 [% of AT1aR mRNA in fraction 2 out of total AT1R mRNA recovered from the sucrose gradient: day 5, 20.9 ± 9.9 vs. day 7, 56.8 ± 5.6, n = 3, P < 0.001]. Accompanying the polysome shift was 50% less RNA-protein complex (RPC) formation between VSMC cytosolic RNA binding proteins in day 7 cells compared with 5-day cultures and the 5′ leader sequence (5′LS) of the AT1aR [5′LS RPC (AU): day 5, 0.62 ± 0.15 vs. day 7, 0.23 ± 0.03; n = 4, P < 0.05] and also with exon 2 [Exon 2 RPC (AU): day 5, 35.0 ± 5.7 vs. day 7, 17.2 ± 3.6; n = 4, P < 0.05]. Taken together, these results suggest that AT1aR expression is regulated by translation during VSMC proliferation in part by RNA binding proteins that interact within exon 2 in the 5′LS of the AT1aR mRNA.
Enhanced Proliferation of Cultured Human Vascular Smooth Muscle Cells Linked to Increased Function of RNA-binding Protein HuR