Abstract
BackgroundProlactin-related proteins (PRPs) are specific proteins of the growth hormone/prolactin (GH/PRL) family in bovine placenta. This study reports the identification and sequencing of a full-length cDNA for two new members of bovine PRPs, bPRP-VIII and -IX, and their localization and quantitative expression in bovine placenta.MethodsNew bPRP-VIII and -IX were identified from bovine placentome. Localization and quantitative gene expression in the placenta were respectively investigated by in situ hybridization and real-time RT-PCR methods. Recombinant proteins of these genes were produced by a mammalian HEK293 cell expression system.ResultsFull-length bPRP-VIII and -IX cDNA were respectively cloned with 909 and 910 nucleotide open-reading-frames corresponding to proteins of 236 and 238 amino acids. The predicted bPRP-VIII amino acid sequence shared about 40 to 70% homology with other bPRPs, and bPRP-IX had about 50 to 80 % homology of others. The two new bPRPs were detected only in the placenta by RT-PCR. mRNA was primarily expressed in the cotyledon and intercotyledonary tissues throughout gestation. An in situ hybridization analysis revealed the presence of bPRP-VIII and -IX mRNA in the trophoblastic binucleate and/or trinucleate cells. bPRP-VIII mRNA was observed in the extra-embryonic membrane on Day 27 of gestation, however, no bPRP-IX mRNA was observed in the extra-embryonic membrane in the same stage of pregnancy by quantitative real-time RT-PCR analysis. Both new bPRP genes were possible to translate a mature protein in a mammalian cell expression system with approximately 28 kDa in bPRP-VIII and 38 kDa in bPRP-IX.ConclusionWe identified the new members of bovine prolactin-related protein, bPRP-VIII and -IX. Localization and quantitative expression were confirmed in bovine placenta by in situ hybridization or real-time PCR. Their different temporal and spatial expressions suggest a different role for these genes in bovine placenta during gestation.
Highlights
Prolactin-related proteins (PRPs) are specific proteins of the growth hormone/ prolactin (GH/PRL) family in bovine placenta
Sequences of bovine PRP (bPRP)-VIII and -IX cDNA Full-length bPRP-VIII and -IX were cloned from bovine placentome
It was shown that bPRP-VIII was close to bPRP-III, bPRP-VI, and bPRP-VII sides in the phylogenetic tree and bPRP-IX was close to bPRP-II and bPRP-IV sides in the phylogenetic tree (Fig. 4)
Summary
Prolactin-related proteins (PRPs) are specific proteins of the growth hormone/ prolactin (GH/PRL) family in bovine placenta. This study reports the identification and sequencing of a full-length cDNA for two new members of bovine PRPs, bPRP-VIII and -IX, and their localization and quantitative expression in bovine placenta. Prolactin-related proteins (PRPs) are members of the growth hormone/prolactin (GH/PRL) family [1]. PRPs are highly expressed in binucleate cells of bovine trophoblasts, their function is still obscure. No new member had been identified between early 1990 and this year [2]. We recently found a new PRP member through a comprehensive analysis using a bovine utero-placental cDNA microarray [3]. Seven bovine PRP (bPRP) genes have been identified in placentomal tissues, whereas only two of those genes have been shown to be translated. BPRP-VII was expressed in intercotyledon and cotyledon with rather weak compare to those of bPRP-I during the gestation [6]
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