Translational regulation of Na,K-ATPase alpha1 and beta1 polypeptide expression in epithelial cells.
To investigate the regulation of the Na,K-ATPase, we have studied the expression of the Na,K-ATPase polypeptides in several mammalian cell lines using the vaccinia virus/T7 RNA polymerase expression system. Infection of several fibroblast-like cell lines with viral recombinants containing the Na,K-ATPase alpha and beta isoforms, the glucose transporters, GLUT 1 and GLUT 4, or the capsid protein of the Sindbis virus all result in the production of the appropriate protein products. However, all epithelial cell lines tested fail to synthesize the Na,K-ATPase viral recombinants, yet they efficiently express the other virally directed polypeptides. While Madin-Darby canine kidney (MDCK) epithelial cells infected with the Na,K-ATPase alpha1 or beta1 recombinant viruses produce both mRNAs, the messages are inefficiently translated. Furthermore, the RNA from infected MDCK cells does not direct the in vitro synthesis of the beta1 polypeptide, whereas the message from infected fibroblast-like BSC 40 cells is efficiently translated both in vivo and in vitro. Moreover, the synthesis of the H,K-ATPase alpha subunit is also limited in MDCK cells, although the H,K-ATPase beta subunit is efficiently expressed. Expression of chimeras constructed between the Na+ pump beta1 isoform and the H,K-ATPase beta subunit indicates that sequences in the 5' coding region of the beta1 message have an inhibitory effect; however, the stringent translational regulation of the beta1 isoform in MDCK cells requires the 5' and 3' regions of the coding sequence. The ability of the polarized cell lines to limit the synthesis of the Na+ pump polypeptides while expressing other vaccinia recombinants at high levels suggests that the polarized cells possess a stringent mechanism for the specific translational regulation of a select set of messages.
Grindstaff
K K
KK
Department of Cell Biology and Physiology, Washington University School of Medicine, St. Louis, Missouri 63110, USA.
Blanco
G
G
Mercer
R W
RW
D000818:Animals; D016764:Cell Polarity; D002478:Cells, Cultured; D004285:Dogs; D004847:Epithelial Cells; D004848:Epithelium / Q000378:metabolism; D005347:Fibroblasts / Q000166:cytology / Q000378:metabolism; D015971:Gene Expression Regulation, Enzymologic; D007668:Kidney / Q000166:cytology / Q000378:metabolism; D014176:Protein Biosynthesis; D011487:Protein Conformation; D012333:RNA, Messenger / Q000032:analysis; D011994:Recombinant Proteins / Q000096:biosynthesis; D012377:Rodentia; D000254:Sodium-Potassium-Exchanging ATPase / Q000096:biosynthesis* / Q000235:genetics; D014616:Vaccinia virus / Q000235:genetics
D012333:RNA, Messenger; D011994:Recombinant Proteins; D000254:Sodium-Potassium-Exchanging ATPase