Identification of band 3-like proteins and Cl-/HCO3- exchange in isolated cardiomyocytes.
The identification of the protein that exerts the function of Cl-/HCO3- exchange is still unresolved in cardiac tissue. We have addressed this issue by using a multiple technical approach. Western blotting analysis with an antibody raised against human erythroid whole band 3 protein, the so-called protein that mediates the Cl-/HCO3- exchange in erythrocytes, showed that adult cardiomyocytes expressed two proteins immunologically related to the erythroid band 3. These proteins migrated in SDS-polyacrylamide gel electrophoresis with apparent molecular masses of 80 and 120 kDa. They were specifically found in the membrane but not in the cytosolic or the myofibril fractions of adult cardiomyocytes. Confocal microscopy further indicated that the immunostained proteins were mainly located at the sarcolemma and along T-tubules, typical membrane structures of adult cardiomyocytes. Using an antibody raised against a cardiac amino-terminal domain of rat AE3, we found that the 120-kDa protein is the translation product of the AE3 gene specifically expressed in heart and brain. Using an antiserum raised against a specific domain of mouse erythroid band 3 (AE1), which is not shared by AE3, we showed that the 80-kDa protein is likely to be a truncated translation product of the AE1 gene. Microinjection of the anti-human erythroid whole band 3 antibody into single isolated cardiac cells significantly inhibited the Cl-/HCO3- exchange activity. Furthermore, the anti-AE1 antibody strongly decreased the efficiency of 4,4'-diisothiocyanatostilbene-2,2'-disulfonate to inhibit the ionic exchange. We thus suggest that the 80-kDa or both the 80- and the 120-kDa proteins immunologically related to the erythroid band 3 protein perform the anionic exchange in rat cardiomyocytes.
Pucéat
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Laboratoire de Physiopathologie Cardiovasculaire, INSERM U-390, Centre Hospitalier Universitaire Arnaud de Villeneuve, Montpellier, France.
Korichneva
I
I
Cassoly
R
R
Vassort
G
G
D000595:Amino Acid Sequence; D000818:Animals; D001457:Anion Exchange Protein 1, Erythrocyte / Q000235:genetics / Q000378:metabolism*; D017920:Antiporters / Q000378:metabolism*; D001639:Bicarbonates / Q000378:metabolism*; D027963:Chloride-Bicarbonate Antiporters; D002712:Chlorides / Q000378:metabolism*; D006801:Humans; D051379:Mice; D008969:Molecular Sequence Data; D009206:Myocardium / Q000166:cytology / Q000378:metabolism*; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D051381:Rats
D001457:Anion Exchange Protein 1, Erythrocyte; D017920:Antiporters; D001639:Bicarbonates; D027963:Chloride-Bicarbonate Antiporters; D002712:Chlorides; D012333:RNA, Messenger