Parathyroid hormone (PTH), acting at least in part through a cAMP signaling pathway, regulates three important transporters in the renal proximal convoluted tubule, namely Na-H exchanger 3, Na-K ATPase, and type IIa sodium phosphate cotransporter (NaPi IIa). The regulation of Na-H exchanger 3 by protein kinase A requires a protein co-factor from the sodium-hydrogen exchanger regulatory factor (NHERF) family of proteins (NHERF-1 and NHERF-2). However, the role of NHERF in PTH regulation of Na-K ATPase and NaPi IIa has not been explored. For studying the role of NHERF-1 on PTH regulation of these transporters, wild-type mNHERF-1 (1-355) or mNHERF-1 (1-325) lacking the ezrin-binding domain were expressed in proximal tubule-derived opossum kidney cells. PTH inhibited Na-K ATPase activity in cells expressing wild-type NHERF-1 associated with increased serine phosphorylation of the alpha subunit of the transporter. By contrast, in cells expressing mNHERF (1-325), the phosphorylation of the alpha subunit of Na-K ATPase was blunted and the activity of the transporter was stimulated in response to PTH. Basal sodium-dependent phosphate transport was lower in cells expressing mNHERF-1 (1-325) as compared with cells expressing mNHERF-1 (1-355). Nonetheless, there were no differences in PTH-associated inhibition of the activity or the decrease in membrane expression of the NaPi IIa in any of the cell lines. These experiments document for the first time an association between NHERF-1 and PTH regulation of Na-K ATPase in epithelial cells. These experiments also suggest that the mechanism for retrieval of NaPi IIa transporters from the apical membrane in response to cAMP does not require NHERF.