Molecular cloning of bovine (Bos taurus) cDNA encoding a 94-kDa glucose-regulated protein and developmental changes in its mRNA and protein content in the mammary gland

We isolated and sequenced cDNA clones encoding a 94-kDa glucose-regulated p rotein (GRP94) from a cDNA library constructed using bovine (Bos taurus) ma mmary gland poly(A) I RNA. The coding nucleotide sequence and the deduced a mino acid sequence of bovine GRP94 shared 94.2-88.4% and 98.1-96.5% identit y with those of other mammalian species, respectively. The primary structur e contained a carboxyl-terminal signal sequence for retention in the endopl asmic reticulum, six potential sites for N-linked glycosylation and two pot ential adenosine 5'-triphosphate binding sites, similar to other mammalian and avian GRP94 homologues. In Northern blot hybridization using a cDNA pro be from the bovine GRP94 cDNA sequence, a transcript 3.0 kb in size was det ected. We measured the amounts of GRP94 and its mRNA in mammary glands from cows at various developmental stages of hormonally induced lactation. The highest level of GRP94 mRNA, determined by dot blot analysis, was detected in the developing stage. In contrast to the mRNA level, the amount of prote in, determined by immunoblot analysis using rabbit antiserum raised against GRP94 purified from bovine brain, was higher in lactating stages than in o thers. The increased level of GRP94 mRNA during the developing stage and th e maintenance of GRP94 protein during lactation suggest that the synthesis of GRP94 is regulated during mammary development and differentiation, and a lso that the protein is involved in a function related to lactation. (C) 20 01 Elsevier Science Inc. All rights reserved.