Expression, localization, and function of an N-terminal half fragment of the rat Na,K-ATPase beta-subunit in HeLa cells.
The N-terminal half of the beta-subunit of rat brain Na,K-ATPase was expressed in HeLa cells transfected with the plasmid pSV2TKneo beta N containing the truncated beta-subunit cDNA to study the assembly and transport of alpha-beta complex. Immunoprecipitation from extracts of metabolically labeled transformed cells demonstrated that the truncated beta-subunit polypeptide (beta N) was neither transported to the plasma membrane nor assembled into an alpha-beta complex with the endogenous alpha-subunit. Cell fractionation experiments showed that the beta N truncated subunit remained unassembled within rough microsomes, suggesting that it never exited from the endoplasmic reticulum (ER). The assembly of the endogenous alpha-and beta-subunits in the beta N-expressing cells was significantly inhibited compared with control cells or with the transformants that did not express the beta N. These results suggest that the N-terminal portion of the beta-subunit interferes with the normal assembly of the endogenous complex which normally takes place in the ER.
Omori
K
K
Department of Physiology, Kansai Medical University, Osaka.
Omori
K
K
Morimoto
T
T
Takada
T
T
Akayama
M
M
Yoshimori
T
T
Sabatini
D D
DD
Tashiro
Y
Y
D000818:Animals; D001692:Biological Transport; D002462:Cell Membrane / Q000378:metabolism; D004247:DNA / Q000737:chemistry; D004721:Endoplasmic Reticulum / Q000378:metabolism; D015870:Gene Expression; D006367:HeLa Cells / Q000201:enzymology; D006801:Humans; D008861:Microsomes / Q000378:metabolism; D011233:Precipitin Tests; D011487:Protein Conformation; D051381:Rats; D000254:Sodium-Potassium-Exchanging ATPase / Q000096:biosynthesis / Q000235:genetics*; D014162:Transfection; D014170:Transformation, Genetic
D004247:DNA; D000254:Sodium-Potassium-Exchanging ATPase