Rat vitamin-D-dependent calcium-binding proteins. Specificity of mRNAs coding for the 7500-Mr protein from duodenum and the 28000-Mr protein from kidney and cerebellum.
mRNA extracted from rat duodenum, kidney and cerebellum was translated in a cell-free reticulocyte lysate system in the presence of L-[35S]methionine. Vitamin-D-dependent calcium-binding proteins (D-CaBPs) were identified by immunoprecipitation using antibodies specific to duodenal D-CaBP (7500 Mr) and cerebellar D-CaBP (28000 Mr). When duodenal mRNA was translated, the immunoprecipitated polypeptide, obtained using antibodies to duodenal D-CaBP, comigrated with the pure small D-CaBP. Only the addition of unlabeled small duodenal D-CaBP prevented the immunoprecipitation of the major protein. Likewise, when mRNA extracted from the kidney and cerebellum was translated, the product immunoprecipitated by antibodies specific to large mammalian D-CaBP was electrophoretically similar to pure 28000-Mr protein, being displaced only by the addition of unlabeled large D-CaBP. The yield of the duodenal D-CaBP synthesized in the reticulocyte lysate assay was remarkably high (about 10%) compared to that of the large D-CaBP with renal (1%) or cerebellar (0.4%) mRNA. In the absence or presence of microsomal membranes, proteins of similar molecular weight were synthesized, suggesting that the biosynthesis of both large and small D-CaBPs do not involve the processing of leader sequences. Moreover in our experimental conditions duodenal poly(A)-rich RNA was unable to direct the synthesis of large D-CaBP while the mRNAs extracted from kidney and cerebellum did not code for the small D-CaBP. Our data indicate that two distinct mRNAs, coding for small and for large vitamin-D-dependent CaBPs, are expressed in specific tissues of the rat.
D000818:Animals; D002135:Calcium-Binding Proteins / Q000302:isolation & purification*; D002531:Cerebellum / Q000378:metabolism*; D004386:Duodenum / Q000378:metabolism*; D005815:Genetic Code; D007120:Immunochemistry; D007668:Kidney / Q000378:metabolism*; D008297:Male; D014176:Protein Biosynthesis; D012333:RNA, Messenger / Q000502:physiology*; D051381:Rats; D011919:Rats, Inbred Strains; D064030:S100 Calcium Binding Protein G / Q000235:genetics / Q000302:isolation & purification*
D002135:Calcium-Binding Proteins; D012333:RNA, Messenger; D064030:S100 Calcium Binding Protein G