THU393 An Internal NPT2A Cryptic PDZ Motif Is A Novel Regulator of PTH- And FGF23-sensitive Phosphate Transport

Disclosure: B.W. Sneddon: None. P.A. Friedman: None. T. Mamonova: None. The Na(+)-dependent NPT2A (SLC34A1) phosphate co-transporter is a major regulator of phosphate homeostasis. Its most notable structural element is a carboxy-terminal PDZ ligand that binds Na(+)/H(+) Exchanger Regulatory Factor-1 (NHERF1, SLC9A3R1), a multidomain PDZ protein that determines apical NPT2A localization and is required for phosphate transport. NPT2A also possesses an uncharacterized internal PDZ ligand. Two recent clinical reports describe congenital hypophosphatemia in children harboring Arg(495)His or Arg(495)Cys variants within the internal PDZ motif. Wild-type internal (494)TRL(496) PDZ binds NHERF1 PDZ2, which we consider a regulatory domain. Ablating the internal PDZ ligand with a (494)AAA(496) variant blocked hormone-sensitive phosphate transport. Complementary approaches, including CRISPR/Cas9 technology, side-directed mutagenesis, confocal microscopy, and modeling, showed that NPT2A Arg(495) His or Arg(495)Cys variants do not support PTH or FGF23 action on phosphate transport. CoIP experiments indicate that both variants bound NHERF1 similarly to WT NPT2A. However, in contrast to WT NPT2A, NPT2A Arg(495)His or Arg(495)Cys variants remain at the apical membrane and are not internalized in response to PTH. We predict that His or Cys substitution of the charged Arg(495) changes the electrostatics, preventing phosphorylation of the upstream Thr(494), interfering with phosphate uptake in response to hormone action, and inhibiting NPT2A trafficking. We advance a model where the carboxy-terminal PDZ ligand defines apical localization of NPT2A, while the internal PDZ ligand is essential for hormone-triggered phosphate transport. Presentation: Thursday, June 15, 2023