Endogenous and cholera toxin-catalyzed ADP-ribosylation of a plasma membrane protein by RL-PR-C cloned rat hepatocytes.
Cholera toxin catalyzed the ADP-ribosylation of a single plasma membrane protein (Mr 55 000) of both RL-PR-C rat hepatocytes and purified rat liver plasma membranes. Labeling of this protein from nicotinamide [2,8-3H]adenine dinucleotide was competitively inhibited by free arginine, but by no other amino acid tested, including lysine. The same protein was ADP-ribosylated from NAD+ endogenously, i.e., in the absence of toxin. This process was, however, not competitively inhibited by added arginine nor by any other amino acid tested lysine. Free ADP-ribose, even in 50-fold molar excess over the nicotinamide [2,8-3H]adenine dinucleotide substrate, did not reduce (by isotope dilution) the endogenous or cholera toxin-catalyzed labeling of the 55 000 dalton membrane protein. It is likely, therefore, that hepatocyte plasma membranes contain an ADP-ribosyltransferase, with a mechanism similar to that of the A subunit of cholera toxin, in that both transfer ADP-ribose to the same membrane protein and in that neither apparently produce free ADP-ribose as an intermediate. It is also clear that the acceptor residue in the 55 000 dalton protein is different for each process. Cholera toxin-catalyzed and endogenous transfer of ADP-ribose to the hepatocyte plasma membrane protein, in contrast to a pigeon erythrocyte system, required no cytosolic factors. The results indicate that ADP-ribosylation in cloned differentiated rat hepatocytes differs from that in pigeon erythrocytes in that the acceptor protein is larger (55 000 compared to 42 000 daltons), cytosolic factors are not required and transfer of ADP-ribose to the acceptor protein occurs endogenously.
D000246:Adenosine Diphosphate Ribose / Q000378:metabolism*; D000262:Adenylyl Cyclases / Q000378:metabolism; D000596:Amino Acids / Q000494:pharmacology; D000818:Animals; D001120:Arginine / Q000494:pharmacology; D002460:Cell Line; D002462:Cell Membrane / Q000187:drug effects / Q000378:metabolism; D002772:Cholera Toxin / Q000494:pharmacology*; D002999:Clone Cells; D007700:Kinetics; D008099:Liver / Q000378:metabolism*; D008565:Membrane Proteins / Q000378:metabolism*; D009243:NAD / Q000378:metabolism; D009702:Nucleoside Diphosphate Sugars / Q000378:metabolism*; D051381:Rats
D000596:Amino Acids; D008565:Membrane Proteins; D009702:Nucleoside Diphosphate Sugars; D009243:NAD; D000246:Adenosine Diphosphate Ribose; D002772:Cholera Toxin; D001120:Arginine; D000262:Adenylyl Cyclases