Submitted on August 14, 2008
Revised on February 13, 2009
Accepted on February 13, 2009
GABARAP deficiency modulates the expression of NaPi-IIa in renal brush border membranes
Sonja C Reining1, Serge M Gisler1, Daniel Fuster2, Orson W Moe MD2, Gregory A O'Sullivan3, Heinrich Betz3, Jürg Biber1, Heini Murer1, and Nati Hernando1*
1 University of Zurich
2 University of Texas Southwestern Med Ctr
3 Max-Planck Inst fur
* To whom correspondence should be addressed. E-mail: hernando{at}physiol.uzh.ch.
Renal reabsorption of inorganic phosphate (Pi) is mainly mediated by the Na-dependent Pi-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 C-terminal PDZ-binding motif (TRL) of NaPi-IIa. In this paper GABARAP was identified as a novel interacting partner of NaPi-IIa using 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 using GST-pull downs from BBM and co-immunoprecipitations 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 Pi, higher Na-dependent 32Pi-uptake in BBMV, and increased expression of NaPi-IIa in renal BBM compared to GABARAP+/+ mice. The expression of NHERF1, 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 (PTH) or acute changes of the dietary Pi content.
