Gene expression for a novel protein RGPR-p117 in various species: the stimulation by intracellular signaling factors

The presence and expression for the gene encoding a novel regucalcin gene promoter region-related protein (RGPR-p117) in various species was investigated by using Southern "zoo blot" and Northern hybridization analyses. A "zoo blot" analysis demonstrated that RGPR-p117 gene was widely conserved in various species including human, rat, mouse, dog, cow, pig, rabbit, chicken, fish, C. elegans and yeast. The gene was not found in Xenopus. Northern blot analysis showed that RGPR-p117 mRNA was expressed in the liver of human, rat, mouse, and rabbit as a single mRNA of approximately 4.5 kb, respectively. However, homologous mRNA was not found in the liver of Xenopus. The expression of RGPR-p117 mRNA in liver was clearly enhanced 5 h after a single intraperitoneal administration of CaCl(2) (5 mg Ca(2+)/100 g body weight) to rats. The RGPR-p117 mRNA is also expressed in the cloned H4-II-E rat hepatoma cells, although this expression was weak as compared with that of liver tissues. Moreover, the RGPR-p117 mRNA expression in H4-II-E cells was stimulated in the presence of dibutyryl cAMP, PMA, insulin, 17beta-estradiol, or serum in culture medium. The present study demonstrates that the RGPR-p117 gene is conserved in various species, and that its expression is stimulated by intracellular signaling factors.     

家蚕磷酸吡哆醇氧化酶基因的表达谱分析

【目的】了解家蚕Bombyx mori维生素B6关键代谢酶磷酸吡哆醇氧化酶（pyridoxine- 5′-phosphate oxidase, PNPO）基因在家蚕不同发育阶段及5龄幼虫不同组织中的表达差异。【方法】将家蚕PNPO基因的重组表达质粒pET-22b(+)-PNPO转化入大肠杆菌Escherichia coli Rosetta中诱导表达, 纯化蛋白制备多克隆抗体。分别采用荧光定量PCR和Western blot方法对家蚕PNPO基因进行了转录水平和翻译水平的表达分析。【结果】在家蚕发育水平上, 5龄幼虫的PNPO翻译量为最高。PNPO基因在5龄幼虫各组织中的转录水平由高到低依次为精巢、 头、 中肠、 马氏管、 卵巢、 表皮、 脂肪体、 丝腺; 翻译量也以精巢为最高, 其次是头、 中肠和马氏管。【结论】明确了PNPO在家蚕各发育阶段及5龄幼虫各组织中的表达情况。     

Structure and expression of the human MEP1A gene encoding the alpha subunit of metalloendopeptidase meprin A

The human genome contains several sequences that are similar to the MEP1A gene encoding the alpha subunit of metalloendopeptidase meprin A. We now report the first genomic structure for the human MEP1A gene that maps to chromosome 6p21. The gene spans approximately 45 kb and consists of 14 exons and 13 introns. Overall, about 6.7% (3 kb) of the MEP1A gene corresponds to the exon sequences. Tissue specificity of the MEP1A gene expression was examined by dot blot analysis of poly(A) RNA from 50 different human tissues. The MEP1A mRNA was detected for the first time in kidney and appendix in addition to colon and small intestine previously known to express the gene. The elucidated gene structure and tissue-specific expression of the MEP1A gene set the stage for investigating regulation and function of the gene and related sequences in the human genome.     

Cloning of a novel member of the reticulon gene family (RTN3): gene structure and chromosomal localization to 11q13.

A novel member of the neuron-specific protein (NSP) or newly named reticulon (RTN) gene family was isolated during a subtraction cloning between macula and peripheral retina. The mRNA for this NSP/RTN-like gene is approximately threefold more abundant in macula than in peripheral retina. The cDNA is 2527 bp long and contains an open reading frame of 236 amino acids. The deduced peptide shows a strong similarity to the NSP/RTN and tropomyosin-like gene families but it is clearly a novel member. The gene contains seven exons and spans more than 15 kb. The gene was localized to chromosome 11q13 between markers D11S4535 and D11S4627 using somatic cell hybrid panels. Southern blot analysis identified the presence of a pseudogene(s) that was subsequently localized to chromosome 4. Multitissue Northern blot analysis found this gene to be widely expressed in human tissues with the highest expression in the brain. We are calling this gene RTN3 to reflect the newly proposed nomenclature.     

Cloning, characterization and genomic organization of LCC-1 (scya16), a novel human CC chemokine expressed in liver

By homology search of expressed sequence tags (EST) in GenBank a novel member of the CC chemokine family was identified. The full-length sequence of this liver-specific CC chemokine (LCC-1) predicted a mature protein of 97 amino acids with 31-48% identity to other CC chemokines. There was a characteristic amino acid C-term extension when aligned with other chemokines. Northern blot analysis from a panel of human tissues revealed that LCC-1 mRNA expression is restricted to adult and fetal liver. Different polyadenylation results in two mRNA species of 1.5 kb and 0.5 kb in size. LCC-1 is constitutively expressed in human HepG2 hepatoma cells and is induced by hypoxic exposure. The promoter region of the LCC-1 gene contains potential HIF-1 binding sites. The EST for LCC-1 has been previously mapped to the CC chemokine cluster on human chromosome 17q11.2. The organization of the LCC-1 gene (scya16) into three exons interrupted by two introns is identical to that found for other members of the CC chemokine family.     

Molecular cloning and characterization of the complementary DNA coding for the B-chain of murine Clq

cDNA clones coding for the B-chain of murine Clq were isolated from a mouse macrophage library. The characterized clones include the total coding region plus a leader sequence. High homology was found with human Clq B-chain in the coding region (81%). Northern blot analysis of total RNA from different tissues of Balb/c mice showed one band of approximately 1.2 kb. The highest signal was found in RNA preparations of thioglycolate-activated peritoneal macrophages. The probe also hybridized with mRNA from spleen, thymus and heart. Extremely weak signals were found in liver, kidney, lung and intestine tissues.