Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts:   Target Concepts:
Query: EC:2.3.1.28 (chloramphenicol acetyltransferase)
 5,100 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The gene encoding the human basal histone variant H2A.Z has been cloned and sequenced. There is a single functional H2A.Z gene with several pseudogene copies. No other histone genes were found in the 3 kilobases of upstream sequence or in the 0.7 kilobase of downstream sequence. In the upstream region, there are regions of Alu sequences, located about 1375 and 2650 base pairs before the transcription start site. The amount of the H2A.Z transcript is unlinked to DNA replication; however, the amount of the H2A.Z transcript is greatly decreased as proliferating cell cultures become quiescent due in part to a decrease in the rate of transcription. Promoter sequences upstream from the H2A.Z gene have been delineated in IMR-90 cells by chloramphenicol acetyltransferase gene expression. Maximal promoter activity was found in a chloramphenicol acetyltransferase construct that contained 234 base pairs just upstream from the transcription start site. This region includes two GC boxes and three CCAAT boxes as well as a properly positioned TATA box. The organization of the human gene is similar to that of the recently characterized chicken gene (Dalton, S., Robins, A. J., Harvey, R. P., and Wells, J. R. E. (1989) Nucleic Acids Res. 17, 1745-1756). Both have four introns with identical exon-intron borders, but three of the introns in the chicken gene are much longer than those in the human. The promoter regions of the two genes have little overall homology; however, two GC boxes and one of the CCAAT boxes are conserved.
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PMID:The human histone H2A.Z gene. Sequence and regulation. 169 87

An 86-kDa heat-shock-protein-encoding (hsp86) cDNA probe permitted to identify, in whole genomic human DNA, two EcoRI fragments of 2.6 and 5.3 kb. These two fragments, as well as an homologous phage lambda VIII1 harboring about 19 kb of human DNA, were isolated from genomic libraries. Sequence analysis revealed that three different genomic hsp86 sequences had been cloned, one of them being the 5' half of a functional gene. This gene contains several introns, as compared to the entire Hsp86-encoding sequence found in lambda VIII1, which represents a processed pseudogene. Cloned hsp86 promoter, with its TATA-box and a heat-shock element upstream at nt positions -25 and -75, respectively, was functional, as verified by fusion to the bacterial chloramphenicol acetyltransferase-encoding gene and its transient expression in vivo. The typical hsp86-type heat-shock regulation was observed, i.e., significant basal activity associated with an inducibility at elevated temperatures. Furthermore, accurate and efficient in vitro transcription was initiated at this hsp86 promoter, resulting in expression of the hsp86 gene, as well as the unrelated sequences.
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PMID:Cloning and analysis of a human 86-kDa heat-shock-protein-encoding gene. 259 42

We report the sequence of the entire human gene encoding beta-glucocerebrosidase and that of the associated pseudogene. The gene contains 11 exons extending from base pair 355 to base pair 7232 in the overall sequence. The gene promoter contains TATA- and CAT-like boxes upstream of the major 5' end of the glucocerebrosidase RNA. The two TATA boxes lie between nucleotides (-23)-(-27) and (-33)-(-39) and the two possible CAT boxes reside between nucleotides (-90)-(-94) and (-96)-(-99) in relation to the major 5' end of the mRNA. The functionality of the promoter region was monitored by coupling it to the bacterial gene coding for chloramphenicol acetyltransferase (CAT) and assaying the expression of the enzyme in cells transfected with this vector. The glucocerebrosidase promoter not only directs synthesis of the bacterial enzyme but also exhibits the same pattern of tissue-specific expression as that of the endogenous gene. An apparently tightly linked pseudogene is approximately 96% homologous to the functional gene. However, introns 2, 4, 6, and 7 have large "deletions" consisting of Alu sequences 313, 626, 320, and 277 bp in length, respectively. It is entirely possible that the ancestral gene lacks these sequences and that they have been inserted into the introns of the functioning gene. There is also a 55-bp deletion from a part of exon 9 flanked by a short inverted repeat. The sequence data should facilitate development of methods for diagnosis of Gaucher disease at the molecular level.
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PMID:The human glucocerebrosidase gene and pseudogene: structure and evolution. 291 9

Genomic clones containing the rat pyruvate kinase M gene, which encodes the M1- and M2-type isozymes, were isolated and their exon sequences were determined. This gene contains 12 exons and 11 introns and is 20 kilobases (kb) long. The sequences specific to the M1- and M2-types exist in exons 9 and 10, respectively (Noguchi, T., Inoue, H., and Tanaka, T. (1986) J. Biol. Chem. 261, 13807-13812). The seventh intron begins with the GC dinucleotide instead of the consensus GT dinucleotide, but other exon-intron boundaries are consistent with the "GT-AG" rule. S1 mapping analysis showed that M1- and M2-type mRNAs had multiple, but the same transcription initiation sites. Thus, the M1- and M2-type isozyme mRNAs are concluded to be produced from the same M gene transcript by alternative RNA splicing. RNA blot hybridization analysis indicated that developmental changes of the isozymes in brain and skeletal muscle were regulated at the level of RNA splicing. The 5'-flanking region of the gene has no "TATA box" or "CAAT box," but contains potential Sp1 binding sites. Bacterial chloramphenicol acetyltransferase assay revealed that a fragment of about 0.5 kb of the 5'-flanking region of the gene was sufficient for promotor activity in the rat hepatoma cell line, dRLh-84. This activity was not present in adult rat hepatocytes, indicating that the 0.5-kb fragment has tissue-specific promoter activity. A processed-type pseudogene that resembles the M2-type pyruvate kinase cDNA was also characterized.
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PMID:Rat pyruvate kinase M gene. Its complete structure and characterization of the 5'-flanking region. 291 12

Cytochrome P4502D6 (CYP2D6) catalyzes the oxidative metabolism of several clinically important classes of drugs. Many of these have lower metabolic clearance rates among Chinese, compared with Caucasians, and are prescribed at lower doses for Asian patients. We have now evaluated the molecular genetic basis for this interethnic difference in drug metabolism. The CYP2D loci from two Chinese subjects, one homozygous for the XbaI 44-kilobase haplotype and one homozygous for the XbaI 29-kilobase haplotype, were cloned and characterized. Sequence analysis revealed two variant CYP2D6 genes, CYP2D6Ch1 and CYP2D6Ch2, having mutations yielding two and eight amino acid substitutions, respectively. Exon 9 of the CYP2D6Ch2 gene contained a sequence of 49 bases originating from the pseudogene CYP2D7P. In addition, mutations in the 5' flanking region common to both CYP2D6Ch genes were found. To evaluate the origin of the detrimental mutation in the genes, parts of the 5' flanking regions were introduced into a Hep G2/simian virus 40 expression system with chloramphenicol acetyltransferase as a reporter gene, and transfected cells were analyzed for activity. The ability of the upstream regions to bind nuclear factors was also evaluated using gel-shift analysis. Furthermore, several chimeric constructs of the CYP2D6wt and CYP2D6Ch genes were made, inserted into pCMV2 vectors, and expressed in COS-1 cells. A part of the upstream region of base pairs -1407 to -1068 was found to constitute an enhancer element, but the CYP2D6Ch-specific mutations did not influence the chloramphenicol acetyltransferase activity in the expression system. In contrast, expression of the chimeric genes revealed that the detrimental mutation of the CYP2D6Ch genes was C188-->T, causing a Pro34-->Ser amino acid substitution in a region that is a highly conserved in cytochromes P450 belonging to gene families 1 and 2. This substitution caused expression of a more unstable gene product, as evident from comparison of the relative levels of CYP2D6 mRNA, CYP2D6 protein, and bufuralol 1'-hydroxylase activities in pCMV2-CYP2D6-transfected COS-1 cells. Allele-specific polymerase chain reaction analysis of genomic DNA from 90 Chinese individuals revealed that the CYP2D6Ch1 allele was the most common one and its distribution correlated well with a higher metabolic ratio for debrisoquine. These data demonstrate that important interethnic differences exist in the structure of the CYP2D locus, and they suggest that the frequent distribution of the C188-->T mutation among the CYP2D6Ch genes explains the lower capacity among Chinese to metabolize drugs that are substrates of CYP2D6, such as antidepressants and neuroleptic agents.
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PMID:Genetic analysis of the Chinese cytochrome P4502D locus: characterization of variant CYP2D6 genes present in subjects with diminished capacity for debrisoquine hydroxylation. 793 25

The gene encoding a folate-binding protein (FBP) expressed in human placenta has been cloned by screening a genomic library with the KB cell FBP complementary DNA. This gene, contained in a 10-kilobase EcoRI fragment of this genomic clone, has 5 exons, 4 introns, the AATAA polyadenylation signal in the 3'-untranslated region, and a 5'-flanking sequence which contains the promoter elements, all of which span approximately 5 kilobases. Transcription initiation was mapped by RNase protection to a site 73 base pairs downstream from a G-rich sequence linked to a tandemly repeated GGAAG sequence which is a motif that the ets oncogene encoded GA-binding protein (GABP) transcription factor binds. Gel-shift and supershift mobility assays indicate that the G-rich sequence and the ets motif bind specifically to SP1 and GABP, respectively. These cis regulatory elements in tandem drive expression of the chloramphenicol acetyltransferase reporter gene in transiently transfected mouse 3T3 cells. The location of these elements upstream of transcription initiation in this gene, which lacks an appropriately located TATA box promoter, indicates that this SP1-GA binding region most probably regulates expression of this placental FBP. The gene encoding this placental FBP has been assigned the FBP/PL-1 gene because it is a member of a multigene family that includes a gene encoding a FBP expressed in both KB cells and placenta and its unprocessed pseudogene.
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PMID:Characterization of the gene encoding a folate-binding protein expressed in human placenta. Identification of promoter activity in a G-rich SP1 site linked with the tandemly repeated GGAAG motif for the ets encoded GA-binding protein. 810 41

A family of 16 genes encoding the mouse ribosomal protein S24 was identified, and four members from this family were cloned. A single expressed intron-containing S24 gene (termed mrpS24) and one pseudogene (mrpS24p) were completely sequenced and characterized. The mrpS24 gene has seven exons and six introns spanning over 5.1 x 10(3) nucleotides (nt). The cap site of S24 was mapped to a G residue four nt upstream of a polypyrimidine tract and 15 nt downstream of a TATA-like (TATGA) element. The 5' region (-325 to +33) of the mrpS24 gene has a functional promoter that was able to express the fused chloramphenicol acetyltransferase (CAT) reporter gene. Two different forms of mouse S24 cDNA clones were previously isolated. Sequence analysis showed that one of these cDNA clones (termed S24a) lacks the entire exon V sequence (18 nt), and the deduced amino acid sequence is missing a C-terminal lysine residue encoded by the other cDNA (S24b). The pseudogene mrpS24p is flanked by an 11-bp direct repeat, and its sequence is almost identical to the S24 cDNA sequence, but it lacks two mini-exons, V and VI (20 nt), as in the cases of the human and rat S24 cDNAs. RT-PCR experiments demonstrated the existence of a third form (S24c) that similarly lacks both of the mini-exons, and suggested that different species of S24 mRNA might arise from alternative splicing of the mini-exons V and VI. Northern blot analysis showed that S24 expression is down- and up-regulated during adipocyte differentiation and in cellular transformation, respectively. RNase protection assays and RT-PCR experiments suggested that these cell-specific changes of S24 mRNA levels are mainly due to fluctuations in S24c mRNA level. Our results provide the first indication that a ribosomal protein gene is regulated by alternative usage of two mini-exons in a cell-specific manner.
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PMID:Molecular characterization of the mouse ribosomal protein S24 multigene family: a uniquely expressed intron-containing gene with cell-specific expression of three alternatively spliced mRNAs. 812 13

We have isolated two non-overlapping clones containing the genes for subunit Vb of cytochrome c oxidase (COXVb) from a rat genomic library in Charon 4A using a newly isolated full-length cDNA as a probe. One of the two genomic clones, designated as lambda COXVb741, contained a functional gene (COXVb-1), while the other one, designated as lambda COXVb211, contained an intronless processed pseudogene (COXVb-2). The COXVb-1 gene spans approximately 1.8 kb and consists of four exons interrupted by three introns. The nucleotide sequences of all exons are completely identical to the corresponding sequences of the rat liver and brain COXVb cDNAs, indicating that this gene is actually expressed. The 5'-flanking region of the gene lacks conventional TATA and CAAT boxes, but exhibits strong promoter activity in the chloramphenicol acetyltransferase (CAT) assay. Deletional analysis and gel shift assay of the 5'-flanking region suggested that the binding of nuclear factor Sp-1 could be essential for transcription of the gene. Southern blotting analysis implied the occurrence of multiple COXVb genes in the rat genome. However, the results of the present experiments suggest that only the COXVb-1 gene is expressed in rat tissues.
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PMID:Molecular cloning and characterization of the rat cytochrome c oxidase subunit Vb gene. 820 67

The pregnancy-specific glycoproteins (PSGs) of the human placenta are a group of proteins that together with the carcinoembryonic antigens comprise a subfamily within the immunoglobulin superfamily. To study the control of PSG expression, we isolated and characterized PSG genes and identified cis-acting DNA elements in the 5'-flanking gene regions essential for PSG expression. Two overlapping PSG cosmid clones, which contain two allelic variants of a PSG gene (PSG12 and PSG12 psi), were isolated from an unamplified library made from a single individual. Cosmid 1 contains exons 1 (5'/L) and 2 (L/N) of the PSG12 gene located downstream of a previously identified PSG1-I gene. Cosmid 6 contains a portion of the PSG1-I gene lacking exons 1 and 2 upstream of a complete PSG12 psi transcription unit. Sequence comparison indicates that exons 5'/L and L/N in PSG12 and PSG12 psi are 99% identical, except that the L/N exon in the PSG12 psi gene contains a stop codon. Both PSG12 and PSG12 psi transcripts were detected in the human placenta, indicating that both genes are actively transcribed. However, the PSG12 psi gene may represent an allelic pseudogene variant of the PSG12 gene, because all identified PSGs contain a functional N-domain. Primer extension analysis showed that the PSG12 gene starts at a cluster of sites located at -106 to -104 base pairs with respect to the translation start site. In transient transfection assays using a chloramphenicol acetyltransferase reporter gene, we demonstrated that the -835 to -34 DNA region upstream of the translation start site of PSG12 or PSG12 psi contained both positive and negative elements that control PSG expression. Deletion analysis showed that nucleotides -172 to -34 in the PSG12 gene could function as a promoter. Gel retardation analysis showed that protein factors in human placental cell extract formed four complexes (I, II, IIa, and III) with the PSG12(-172/-34) DNA. Site-directed mutagenesis that prevents protein factor binding to the PSG12 promoter resulted in a marked reduction in transcription activation, locating the core enhancers at nucleotides -148 to -141 and -60 to -55. Mutagenesis studies also showed that the ACAGC repeats at nucleotides -84 to -68 in the PSG12 5'-flanking are essential for expression of the PSG12 gene in human placental cells.
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PMID:Characterization of two allelic variants of a human pregnancy-specific glycoprotein gene. 834 32

We isolated and characterized four forms of rat CYP11B genes, which were tentatively named CYP11B1, -B2, -B3, and -B4. Genomic Southern analyses indicated that the members of the rat CYP11B gene subfamily were confined to these four genes; among them, CYP11B1 and -B2 encoded steroid 11 beta-hydroxylase and aldosterone synthase, respectively, while CYP11B3 was a gene highly homologous to CYP11B1 without a known expression product. By being devoid of a region spanning two exons conserved in the other three, CYP11B4 was presumably a pseudogene. In the nucleotide sequences, CYP11B1, -B3, and -B4 showed 95-96 and 93-100% identities in the coding and 0.5-kilobase 5'-flanking regions, respectively. However, the homology between the nucleotide sequences of one of the three and CYP11B2 was rather low, about 90 and 50% in the coding and 0.5-kilobase 5'-flanking regions, respectively. As a whole, CYP11B2 rather than CYP11B1, -B3, or -B4 was more homologous to CYP11B genes of other animals such as cow and human. In transient transfection experiments using mouse adrenocortical Y1 cells and chloramphenicol acetyltransferase gene constructs, the 0.5-kilobase 5'-flanking region of CYP11B1 had a 4- and 10-fold higher promoter activity than the corresponding regions of CYP11B2 and -B3, respectively. The possible presence of a suppressive element(s) was noted in the upstream of the 0.5-kilobase region of CYP11B1. Although a variant of cAMP-responsive element, which was present in rat CYP11B2 and all known CYP11B genes of other animals, was modified in rat CYP11B1 and -B3 genes, dibutyryl cAMP stimulated all the promoter activities of the 5'-flanking regions of the rat genes by 3-fold.
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PMID:Isolation and characterization of rat CYP11B genes involved in late steps of mineralo- and glucocorticoid syntheses. 847 52


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