title: GABARAP deficiency modulates expression of NaPi-IIa in renal brush-border membranes.
abstract: Renal reabsorption of inorganic phosphate (P(i)) is mainly mediated by the Na(+)-dependent P(i)-cotransporter NaPi-IIa that is expressed in the brush-border membrane (BBM) of renal proximal tubules. Regulation and apical expression of NaPi-IIa are known to depend on a network of interacting proteins. Most of the interacting partners identified so far associate with the COOH-terminal PDZ-binding motif (TRL) of NaPi-IIa. In this study GABA(A) receptor-associated protein (GABARAP) was identified as a novel interacting partner of NaPi-IIa applying a membrane yeast-two-hybrid system (MYTH 2.0) to screen a mouse kidney library with the TRL-truncated cotransporter as bait. GABARAP mRNA and protein are present in renal tubules, and the interaction of NaPi-IIa and GABARAP was confirmed by using glutathione S-transferase pulldowns from BBM and coimmunoprecipitations from transfected HEK293 cells. Amino acids 36-68 of GABARAP were identified as the determinant for the described interaction. The in vivo effects of this interaction were studied in a murine model. GABARAP(-/-) mice have reduced urinary excretion of P(i), higher Na(+)-dependent (32)P(i) uptake in BBM vesicles, and increased expression of NaPi-IIa in renal BBM compared with GABARAP(+/+) mice. The expression of Na(+)/H(+) exchanger regulatory factor (NHERF)1, an important scaffold for the apical expression of NaPi-IIa, is also increased in GABARAP(-/-) mice. The absence of GABARAP does not interfere with the regulation of the cotransporter by either parathyroid hormone or acute changes of dietary P(i) content.
authors.affiliation: Institute of Physiology and Zurich Center for Integrative Human Physiology (ZIHP University of Zurich, Winterthurerstr. 190, 8057 Zurich, Switzerland.
authors.forename: Sonja C
authors.initials: SC
authors.lastname: Reining
authors.forename: Serge M
authors.initials: SM
authors.lastname: Gisler
authors.forename: Daniel
authors.initials: D
authors.lastname: Fuster
authors.forename: Orson W
authors.initials: OW
authors.lastname: Moe
authors.forename: Gregory A
authors.initials: GA
authors.lastname: O'Sullivan
authors.forename: Heinrich
authors.initials: H
authors.lastname: Betz
authors.forename: Jürg
authors.initials: J
authors.lastname: Biber
authors.forename: Heini
authors.initials: H
authors.lastname: Murer
authors.forename: Nati
authors.initials: N
authors.lastname: Hernando
mesh_terms: D000818:Animals; D051017:Apoptosis Regulatory Proteins; D002460:Cell Line; D003598:Cytoskeletal Proteins / Q000235:genetics* / Q000378:metabolism*; D053595:Embryonic Stem Cells / Q000502:physiology; D004705:Endocytosis / Q000187:drug effects / Q000502:physiology; D004847:Epithelial Cells / Q000502:physiology; D015870:Gene Expression / Q000502:physiology; D015723:Gene Library; D006706:Homeostasis / Q000502:physiology; D006801:Humans; D007668:Kidney / Q000166:cytology; D007687:Kidney Tubules, Proximal / Q000166:cytology / Q000502:physiology*; D008565:Membrane Proteins / Q000235:genetics* / Q000378:metabolism*; D051379:Mice; D008817:Mice, Mutant Strains; D008869:Microtubule-Associated Proteins; D008871:Microvilli / Q000502:physiology; D010281:Parathyroid Hormone / Q000494:pharmacology; D010710:Phosphates / Q000378:metabolism / Q000494:pharmacology; D010750:Phosphoproteins / Q000235:genetics / Q000378:metabolism; D016226:Phosphorus, Dietary / Q000494:pharmacology; D012333:RNA, Messenger / Q000378:metabolism; D017923:Sodium-Hydrogen Exchangers / Q000235:genetics / Q000378:metabolism; D050613:Sodium-Phosphate Cotransporter Proteins, Type IIa / Q000235:genetics* / Q000378:metabolism*
chemical_list: D051017:Apoptosis Regulatory Proteins; D003598:Cytoskeletal Proteins; C480861:GABARAP protein, mouse; D008565:Membrane Proteins; D008869:Microtubule-Associated Proteins; D010281:Parathyroid Hormone; D010710:Phosphates; D010750:Phosphoproteins; D016226:Phosphorus, Dietary; D012333:RNA, Messenger; C493555:Slc34a1 protein, mouse; D017923:Sodium-Hydrogen Exchangers; D050613:Sodium-Phosphate Cotransporter Proteins, Type IIa; C093376:sodium-hydrogen exchanger regulatory factor