Tmp21-I, A VESICULAR TRAFFICKING PROTEIN, IS DIFFERENTIALLY EXPRESSED DURING INDUCTION OF THE URETER AND METANEPHROS

Abstract

To identify genes participating in the reciprocal induction of the metanephros and ureter. Embryonic day 14 Sprague-Dawley rat kidneys and ureters were microdissected into differentiating mesenchyme, ureteric buds, and extrarenal ureter and prepared for RT/PCR differential display. Differentially displayed cDNAs were reamplified, cloned, and sequenced. Expression was verified in the embryonic, newborn or adult kidneys by Northern blot hybridization or RT/PCR using sequence specific primers. A newborn rat kidney cDNA library was prepared and screened with probes of interest. Positive clones were screened, sequenced and compared to the GenBank/EMBL databases. A rabbit polyclonal antibody was raised to a synthetic peptide of the Tmp21-I protein and was used for immunohistochemistry. From the cDNAs differentially displayed by the ureteric buds cDNA B11, is 254 bp in length. The gene for B11 is expressed in adult and newborn kidneys as two transcripts (3.4 kb and 1.3 kb). More importantly, RT/PCR on E14 kidneys using B11 sequence specific primers identified expression in the embryonic kidney at the beginning of induction. B11 cDNA library screening yielded clones with inserts of 1.3 kb. This sequence encodes Tmp21-I, a vesicular trafficking protein. Immunohistochemistry demonstrates that Tmp21-I is abundant in the nephrogenic cortex of the newborn kidney and as a nephron matures, the protein levels decline. The protein is essentially absent in the adult rat kidney. Tmp21-I is a developmentally regulated gene expressed during kidney induction. Localized within the nephrogenic zone, it may direct the intracellular trafficking or secretion of proteins responsible for nephrogenesis.