P(i) transport, phosphorylation, and dephosphorylation in renal membranes from HYP/Y mice

To ascertain whether cAMP-dependent phosphorylation could be demonstrated in brush border membrane vesicles (BBMV) isolated from kidneys of mice with X-linked hypophosphatemic rickets (HYP/Y) and normal littermates (+/Y) and, if so, to determine whether the absence of dephosphorylation might underlie differences in Na⁺-dependent ³²P(i) transport in BBMV, we measured 1) ³²P(i) transport, 2) cAMP-dependent phosphorylation, and 3) dephosphorylation in BBMV from +/Y and HYP/Y mice. Na⁺ gradient-dependent ³²P(i) transport was decreased in BBMV from HYP/Y mice as reflected in a decreased apparent V(max). cAMP-dependent phosphorylation of a 62,000 M(r) protein was demonstrated in sodium dodecyl sulfate polyacrylamide gels of BBMV from +/Y and HYP/Y mice and was associated with decreased Na⁺-dependent ³²P(i) transport. Dephosphorylation of the 62,000 M(r) band was demonstrable in both types of membranes. Thus, both cAMP-dependent protein kinase and phosphoprotein phosphatase activities were demonstrable in BBMV isolated from +/Y and HYP/Y mice. These results are consistent with the renal tubular defect in the HYP/Y mouse reflecting an intrinsic abnormality of P(i) transport in the brush border membrane independent from mediation of the phosphaturic effect of parathyroid hormone.