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 th is issue by using a multiple technical approach, Western blotting anal ysis with an antibody raised against human erythroid whole band 3 prot ein, the so-called protein that mediates the Cl-/HCO3- exchange in ery throcytes, showed that adult cardiomyocytes expressed two proteins imm unologically related to the erythroid band 3, These proteins migrated in SDS-polyacrylamide gel electrophoresis with apparent molecular mass es of 80 and 120 kDa, They were specifically found in the membrane but not in the cytosolic or the myofibril fractions of adult cardiomyocyt es. Confocal microscopy further indicated that the immunostained prote ins 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 th e 120-kDa protein is the translation product of the AE3 gene specifica lly expressed in heart and brain, Using an antiserum raised against a specific domain of mouse erythroid band 3 (AE1), which is not shared b y AE3, we showed that the 80-kDa protein is likely to be a truncated t ranslation product of the AE1 gene, Microinjection of the anti-human e rythroid whole band 3 antibody into single isolated cardiac cells sign ificantly inhibited the Cl-/HCO3-, exchange activity, Furthermore, the anti-AE1 antibody strongly decreased the efficiency of 4,4'-diisothio cyanatostiibene-2,2'-disulfonate to inhibit the ionic exchange, We thu s suggest that the 80-kDa or both the 80 and the 120-kDa proteins immu nologically related to the erythroid band 3 protein perform the anioni c exchange in rat cardiomyocytes.