Protein-RNA interactions determine the stability of the renal NaPi-2 cotransporter mRNA and its translation in hypophosphatemic rats

Hypophosphatemia leads to an increase in type II Na+ dependent inorganic ph osphate cotransporter (NaPi-2) mRNA and protein levels in the kidney and in creases renal phosphate reabsorption. Nuclear transcript run-on experiments showed that the effect of a low phosphate diet was post-transcriptional. I n an in vitro degradation assay, renal proteins from hypophosphatemic rats stabilized the NaPi-2 transcript g-fold compared with control rats and this was dependent upon an intact NaPi-2 3'-untranslated region (UTR), To deter mine an effect of hypophosphatemia upon NaPi-a protein synthesis, the incor poration of injected [S-35]methionine into renal proteins was studied in vi vo, Hypophosphatemia leads to increased [S-35]methionine incorporation only into NaPi-2 protein. The effect of hypophosphatemia on translation was stu died in an in vitro translation assay, where hypophosphatemic renal protein s led to increased translation of NaPi-2 and other transcripts. NaPi-2 RNA interaction with cytosolic proteins was studied by UV cross-linking and Nor thwestern gels. Hypophosphatemic proteins led to increased binding of renal cytosolic proteins to the 5'-UTR of NaPi-2 mRNA. Therefore, hypophosphatem ia increases NaPi-2 gene expression post-transcriptionally, which correlate s with a more stable transcript mediated by the 3'-UTR, and an increase in NaPi-2 translation involving protein binding to the 5'-UTR. These findings show that phosphate regulates gene expression by affecting protein-RNA inte ractions in vivo.