Phosphate (P_i)-regulated heterodimerization of the high-affinity sodium-dependent P_i transporters PiT1/Slc20a1 and PiT2/Slc20a2 underlies extracellular P_i sensing independently of P_i uptake.

ABSTRACT: Extracellular phosphate (P_i) can act as a signaling molecule that directly alters gene expression and cellular physiology. The ability of cells or organisms to detect changes in extracellular P_i levels implies the existence of a P_i-sensing mechanism that signals to the body or individual cell. However, unlike in prokaryotes, yeasts, and plants, the molecular players involved in P_i sensing in mammals remain unknown. In this study, we investigated the involvement of the high-affinity, sodium-dependent P_i transporters PiT1 and PiT2 in mediating P_i signaling in skeletal cells. We found that deletion of PiT1 or PiT2 blunted the P_i-dependent ERK1/2-mediated phosphorylation and subsequent gene up-regulation of the mineralization inhibitors matrix Gla protein and osteopontin. This result suggested that both PiTs are necessary for P_i signaling. Moreover, the ERK1/2 phosphorylation could be rescued by overexpressing P_i transport-deficient PiT mutants. Using cross-linking and bioluminescence resonance energy transfer approaches, we found that PiT1 and PiT2 form high-abundance homodimers and P_i-regulated low-abundance heterodimers. Interestingly, in the absence of sodium-dependent P_i transport activity, the PiT1-PiT2 heterodimerization was still regulated by extracellular P_i levels. Of note, when two putative P_i-binding residues, Ser-128 (in PiT1) and Ser-113 (in PiT2), were substituted with alanine, the PiT1-PiT2 heterodimerization was no longer regulated by extracellular P_i These observations suggested that P_i binding rather than P_i uptake may be the key factor in mediating P_i signaling through the PiT proteins. Taken together, these results demonstrate that P_i-regulated PiT1-PiT2 heterodimerization mediates P_i sensing independently of P_i uptake.

SUBMITTER: Bon N 

PROVIDER: S-EPMC5808770 | biostudies-literature | 2018 Feb

REPOSITORIES: biostudies-literature