Control of Na -K -ATPase 1-subunit expression: role of 3 -untranslated region

Shao, Yvonne, and Faramarz Ismail-Beigi. Control of Na /K ATPase 1-subunit expression: role of 3 -untranslated region. Am J Physiol Cell Physiol 286: C580–C585, 2004; 10.1152/ajpcell.00117. 2003.—Using in vitro translation and cell transfection assays, we previously demonstrated that the Na -K -ATPase 1 mRNA species containing its longest 3 -untranslated region (UTR) exhibited the lowest translational efficiency. Here, employing deletions and in vivo expression assays, using direct injection of plasmids into rat ventricular myocardium, we identified a 143-nt segment located in the distal 3 -UTR of 1 mRNA that was associated with decreased luciferase expression; interestingly, this segment contains three AUUUA motifs. Using RNA-protein binding assays and UV cross-linking of cRNA with cytosolic proteins of rat heart, we identified an 38-kDa protein that specifically bound to the cRNA encoding the 143-nt segment of 1 mRNA 3 -UTR. Mutation of three nucleotides located in the middle region of the 143-nt segment, which was predicted to greatly disrupt a putative stem-loop structure of the cRNA in this region, was associated with reduced binding of the mutated cRNA to the protein migrating at 38 kDa. The cRNA encoding a segment of cyclooxygenase-2 mRNA 3 -UTR containing six AUUUA sequences did not bind the protein migrating at 38 kDa and did not compete with the binding of the wild-type 143-nt 1 cRNA to the protein. The above results suggest that the 143-nt segment in the distal segment of the 3 -UTR of 1 mRNA may play an important role in the control of 1-subunit expression.