title: Pit2 assemblies at the cell surface are modulated by extracellular inorganic phosphate concentration.
abstract: Pit2 is a type III sodium-dependent phosphate transporter and the cell surface receptor for amphotropic murine leukemia virus. Indirect arguments have previously suggested that retrovirus receptor assembly play a role in triggering membrane fusion. Using CHO cells expressing physiological amounts of functional versions of human Pit2 fused to various tagging epitopes, we provide evidence that Pit2 forms assemblies at the cell surface. Living cells were exposed to cross-linking reagents and protein extracts were treated with trifluoroacetic acid (TFA), a chemical that destroys all protein interactions but covalent links. Assemblies were also detected in the absence of cross-linking and TFA treatment, indicating that they are partially resistant to detergent denaturation. The formation of homo-oligomers was documented by the coimmunoprecipitation of differently tagged molecules. The amounts of Pit2 assemblies detected in the presence or in the absence of cross-linking reagents varied with extracellular inorganic phosphate concentration ([P(i)]). Variation of signal intensity was in the range of twofold, occurred in the absence of de novo protein synthesis and took place at the cell surface. These results indicate that Pit2 assemblies exhibit variable conformations at the surface of living cells. Susceptibility to virus infection and phosphate uptake also vary with extracellular [P(i)]. A model is proposed in which cell surface Pit2 assemblies switch from a compacted to an expanded configuration in response to changes of extracellular [P(i)], and possible relationships with the variation of biological activities are discussed.
authors.affiliation: Unité Rétrovirus et Transfert Génétique, CNRS URA 1930, Institut Pasteur, 75724 Paris, France.
authors.forename: Christine
authors.initials: C
authors.lastname: Salaün
authors.forename: Emmanuel
authors.initials: E
authors.lastname: Gyan
authors.forename: Pierre
authors.initials: P
authors.lastname: Rodrigues
authors.forename: Jean Michel
authors.initials: JM
authors.lastname: Heard
mesh_terms: D000818:Animals; D016466:CHO Cells; D006224:Cricetinae; D003432:Cross-Linking Reagents / Q000494:pharmacology; D006801:Humans; D010710:Phosphates / Q000494:pharmacology*; D011233:Precipitin Tests; D011991:Receptors, Virus / Q000378:metabolism*; D012190:Retroviridae / Q000235:genetics / Q000472:pathogenicity; D050608:Sodium-Phosphate Cotransporter Proteins; D050611:Sodium-Phosphate Cotransporter Proteins, Type III; D027981:Symporters / Q000187:drug effects* / Q000378:metabolism*; D014162:Transfection
chemical_list: D003432:Cross-Linking Reagents; D010710:Phosphates; D011991:Receptors, Virus; C493589:SLC20A2 protein, human; D050608:Sodium-Phosphate Cotransporter Proteins; D050611:Sodium-Phosphate Cotransporter Proteins, Type III; D027981:Symporters