Overexpression of the erythrocyte plasma membrane Ca2+ pump in COS-1 cells.
A full-length cDNA corresponding to the hPMCA4 plasma membrane Ca2+ pump was assembled and expressed in COS-1 cells. The original sequence of hPMCA4 gave a very low expression. The mutation of the initiation translation site of this sequence to the consensus A/G-X-X-AUG-G increased the production of the protein. The Ca2+ pump activity in transfected cells was 1.5-3.5-fold higher than in controls. The Ca(2+)-dependence and the calmodulin stimulation of hPMCA4 expressed in COS-1 cells were comparable with those of the erythrocyte Ca2+ pump. Immunohistochemistry experiments showed that most of the expressed protein remained in intracellular membranes. Possible explanations for this targeting of the pump are discussed.
Adamo
H P
HP
Mayo Foundation, Department of Biochemistry and Molecular Biology, Rochester, MN 55905.
Verma
A K
AK
Sanders
M A
MA
Heim
R
R
Salisbury
J L
JL
Wieben
E D
ED
Penniston
J T
JT
D000818:Animals; D001483:Base Sequence; D015153:Blotting, Western; D002118:Calcium / Q000378:metabolism / Q000494:pharmacology; D000252:Calcium-Transporting ATPases / Q000235:genetics* / Q000378:metabolism / Q000502:physiology; D002460:Cell Line; D004247:DNA / Q000737:chemistry / Q000235:genetics; D004910:Erythrocyte Membrane / Q000737:chemistry*; D005455:Fluorescent Antibody Technique; D015870:Gene Expression; D006801:Humans; D008969:Molecular Sequence Data; D014162:Transfection
D004247:DNA; D000252:Calcium-Transporting ATPases; D002118:Calcium