Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts:   Target Concepts:
Query: EC:2.7.7.6 (RNA polymerase)
 34,946 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The sites of expression in the small intestine and the function of CYP2J4, a recently identified rat cytochrome (P450) isoform found to be predominantly expressed in the small intestine, were characterized. Immunoblot analysis with a polyclonal antibody to heterologously expressed CYP2J4 revealed that expression of CYP2J4 was at the highest level in the distal duodenum and jejunum and decreased toward the ileum. Villous cells expressed higher levels of CYP2J4 than crypt cells. Isoform-specific RNA polymerase chain reaction indicated that a related P450 isoform, CYP2J3, was only a minor form in rat small intestine. Since the intestinal mucosa is exposed to high levels of dietary nutrients, we hypothesized that CYP2J4 may be active toward diet-derived factors. We determined that purified, heterologously expressed CYP2J4 is active toward all-trans- and 9-cis-retinal in reconstituted systems, producing the corresponding retinoic acids as the major products. Apparent K(m) values for the formation of retinoic acids were 54 and 49 microM, respectively, and apparent Vmax values were 20 and 21 nmol/min/nmol P450, respectively. These activities were readily inhibited by a polyclonal anti-CYP2J4 antibody. Rat enterocyte microsomes were also active with all-trans-retinal to produce all-trans-retinoic acid in the presence of NADPH, and the majority of retinoic acid synthesis activity was inhibited by the polyclonal anti-CYP2J4 antibody. These findings suggest that CYP2J4 plays a major role in intestinal microsomal metabolism of retinal to retinoic acid and may be involved in the maintenance of retinoid homeostasis in the small intestine in vivo.
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PMID:Characterization of the cytochrome P450 CYP2J4: expression in rat small intestine and role in retinoic acid biotransformation from retinal. 960 60

Expressed sequence tag (EST) and digital Northern analyses of human fetal, adult, and hypertrophic heart cDNA libraries revealed ESTs with high homology to adenomatosis polyposis coli (APC) and its associated protein, beta-catenin, as well as their differential expression. Thus, we hypothesize that the APC/beta-catenin pathway may play a role in cardiac development and disease. Reverse transcriptase-polymerase chain reaction analysis exhibited a higher APC expression in adult compared with fetal and hypertrophic heart but no significant difference in beta-catenin mRNA level. However, beta-catenin protein level was higher in fetal and hypertrophic heart compared with adult heart, suggesting the post-translational regulation of beta-catenin by APC in the cardiovascular system. In vitro antisense inhibition of APC resulted a higher beta-catenin protein expression leading to an incomplete myotube formation, suggesting APC/beta-catenin pathway involvement in myotube development. Western blot analysis further reveals three novel isoforms, APC-F, APC-A, and APC-D, ubiquitously expressed in fetal, adult, and hypertrophic heart, respectively. Isoform switching during development and disease pathogenesis suggests functionally distinct roles for each isoform. These data (i) demonstrate the usefulness of genome-based expression analysis for rapid discovery of differentially expressed genes, (ii) implicate the APC/beta-catenin pathway in the cardiovascular development, and (iii) demonstrate APC isoform switching during cardiac development and disease.
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PMID:Role of the adenomatous polyposis coli gene product in human cardiac development and disease. 1074 62

The Ca2+ release channel of the sarcoplasmic reticulum (SR) is essential for the release of Ca2+ from intracellular stores and is expressed widely in various excitable cells. It plays a key role particularly in excitation contraction coupling in myocytes in skeletal and cardiac muscle. Three isoforms of the SR Ca2+ release channel have been cloned. Recently coexpression of different isoforms was reported in different animal species and various tissues. In human cardiac tissue, however, isoform expression is not yet established. Therefore the aim of this study was to characterize isoform expression of the SR Ca2+ release channel in the human heart. We examined specific isoform expression of mRNA and proteins of the SR Ca2+ release channel in the four different chambers of the heart and the interventricular septum from explanted human hearts from nonfailing organ donors (n=8). Reverse transcriptase PCR from total cardiac RNA with isoform specific primers and western blots from myocardial homogenates with isoform specific antibodies were performed. Quantification of protein expression was achieved by densitometric scanning and computer analysis and is expressed as densitometric units per microgram of protein. A single band DNA signal was detected by reverse transcriptase PCR for the skeletal isoform 1 and the cardiac isoform 2 and isoform 3 in all regions of the human heart investigated. Specific protein expression was detected in all five myocardial regions of the human heart in western blots for the skeletal isoform I and cardiac isoform 2, and a weaker specific band was also detectable for isoform 3 of the SR Ca2+ release channel. Quantification of protein expression showed significant (P=0.008) lower expression of isoform 1 in the right ventricle (42+/-4 densitometric units/g tissue) and similar expression in all other regions (right atrium 58+/-3; septum 51+/-5, left atrium 54+/-5; left ventricle 51+/-6). Isoform 2 of the SR Ca2+ release channel was also significantly lower (P=0.001) in the right ventricle (33+/-4 densitometric/g tissue) and similar in the other heart chambers (right atrium 42+/-5: septum 41+/-3, left atrium 52+/-6, left ventricle 42+/-3). Differences in isoform 3 of the SR Ca2+ release channel for the various myocardial regions did not reach significant levels (right atrium 45+/-6, right ventricle 38+/-5, septum 49+/-8, left atrium 46+/-7, and in left ventricle 45+/-3 densitometric units/g tissue). In conclusion, all three isoforms of the SR Ca2+ release channel were determined in the human heart at both mRNA and protein levels with different quantitative expression in the different heart chambers. Coexpression of the three different isoforms with different functional properties might increase the complexity of regulation of excitation contraction coupling in the human heart in a chamber specific mode.
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PMID:Isoform expression of the sarcoplasmic reticulum Ca2+ release channel (ryanodine channel) in human myocardium. 1100 33

Endothelin-converting enzyme (ECE)-1 is a membrane-bound metalloprotease responsible for production of vasoactive endothelin (ET)-1 from inactive big ET-1. ECE-1 exists as four separate isoforms, ECE-1a, b, c, and d, which differ only in their amino-terminal regions. We investigated the expression and localization of the ECE-1 isoforms in primary human umbilical vein endothelial cells (HUVECs) and EAhy926 cells. Reverse transcriptase polymerase chain reaction showed expression of all four isoforms in both cell lines, with ECE-1d seeming, at least qualitatively, to be the predominant isoenzyme. Isoform-specific polyclonal antibodies were used to investigate isoform protein expression. ECE-1a, b, and c protein was detected in EAhy926 cells by immunoblotting; only ECE-1a and ECE-1c were detected in HUVECs. Using immunofluorescence microscopy analysis, both HUVEC and EAhy926 cells showed nuclear immunoreactivity with a monoclonal antibody recognizing all ECE-1 isoforms. The ECE-1a antibody also showed nuclear immunoreactivity in both cell lines; this seemed to colocalize with nucleolin. The ECE-1b antibody showed nuclear immunoreactivity in EAhy926 cells, but no overlap with nucleolin was seen. Intracellular immunoreactivity was seen in both cell lines using the ECE-1c antibody; this showed some colocalization with concanavalin A (an endoplasmic reticulum marker). von Willebrand Factor was used as a marker for Weibel-Palade bodies in HUVECs, but no colocalization with ECE-1 was seen during this study. The data presented here sheds new light on the localization of ECE-1a, b, and c in cultured human endothelial cells, which may further understanding of the ET system and aid design of therapeutic ECE inhibitors.
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PMID:Expression and localization of endothelin-converting enzyme-1 isoforms in human endothelial cells. 1674 Sep 87