PHOSPHATE DEPRIVATION INDUCES OVEREXPRESSION OF 2 PROTEINS RELATED TOTHE RAT RENAL PHOSPHATE COTRANSPORTER NAPI-2

Polyclonal antibodies were raised in rabbits against the C-terminal po rtion of the rat renal brush-border membrane sodium/phosphate cotransp orter NaPi-2. Antibody specificity and molecular sizes of proteins rel ated to NaPi-2 were assayed by Western blot analysis. Proteins of 40 a nd 70-75 kDa (p40 and p70) were immunodetected in rat and mouse brush- border membranes and proteins of 72 and 82 kDa were detected in rabbit . The absence or presence of beta-EtSH in the samples before electroph oresis greatly influenced the immunodetection profile of the rat prote ins. Since the 40 kDa protein (p40) can only be detected under reducin g conditions, it probably originates from reduction of disulfide bonds in p70. Tryptic cleavage of p40 and p70 revealed identical protein fr agments showing the close structural identity of those proteins. Both proteins were more abundant in the outer cortex portion of the rat kid ney than in the juxtamedullary portion. Furthermore, rats fed a low-ph osphate diet for 24 h showed a 20- and 14-fold increase in the amount of p40 and p70, respectively, compared to control rats, showing that t he adaptation to P-i deprivation by increasing renal phosphate reabsor ption is not only the result of overproduction of p70, as previously s hown, but is also due to the novel p40 which most probably derives fro m p70.