Mechanism of renal phosphate retention during growth

Abstract

We have previously demonstrated that the retention of phosphate required for growth is due to a a high Vmax of the Na(+)-Pi cotransport system located in the brush border membrane of the proximal tubule. Because of this and other similarities between adaptation of the kidney to a high Pi demand (growth) and that to low Pi supply, we measured the levels of NaPi-2 mRNA and cDNA present in kidney cortex of 3- and > 12-week-old rats. Like in Pi depletion, Western blots revealed that a 80 to 85 kDa protein recognized by a polyclonal antibody directed against the N-terminal region of the NaPi-2 protein was 2.3-fold more abundant in renal microvilli of the young than of adult animals. However, unlike in Pi depletion, Northern blot analysis failed to reveal a significant difference between mRNA levels at the two ages. Furthermore, suppression of NaPi-2 mRNA activity by annealing with antisense oligomers, or removal of the NaPi-2 transcripts by subtractive hybridization did not affect the rate of Na(+)-Pi cotransport induced in oocytes by polyA RNA of rapidly growing animals, while abolishing the ability of the renal cortical polyA RNA of adult rats to encode for Na(+)-Pi cotransport. RT-PCR of subtracted polyA RNA using primers specific for a region conserved in NaPi type II (Pi modulated) cotransporters yielded a product that was 98% homologous with that region, despite the absence of NaPi-2 cDNA. The results of these experiments demonstrate that the polyA RNA from kidneys of young animals contains unique mRNA transcripts able to encode for a NaPi protein homologous to, but distinct from NaPi-2.