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R S Seelan

Publications and source records attributed to R S Seelan.

17 recordsLinked to original sources
PubMed2001

Genomic structure, chromosome mapping and expression analysis of the human AXIN2 gene.

Conductin is a Wnt signalling protein and serves as a negative regulator of beta-catenin stability. We have previously isolated the human homolog (AXIN2) of the murine conductin gene and shown that it is mutated in colorectal cancer (CRC) with defective mismatch repair (MMR). Here we report the detailed genomic structure of this gene by analysis of cDNA and genomic clones. The gene spans > or =25 kb containing ten exons ranging from 96 bp to 904 bp. All splice donor and acceptor sites conform to the GT/AG rule. FISH (Fluorescence in situ Hybridization) analysis localized this gene to human chromosome band 17q24 and showed that it exists as a single copy in the human genome. Northern blot analysis from different human organs demonstrated that the AXIN2 gene is highly expressed in human thymus, prostate, testis, small intestine and ovarian tissues but expressed at a lower level in colon. The data reported here provides a framework for further analysis of this important Wnt signalling protein in vertebrate development and tumorigenesis.

Axin Protein↗
PubMed2000

Role for the p53 homologue p73 in E2F-1-induced apoptosis.

The transcription factor E2F-1 induces both cell-cycle progression and, in certain settings, apoptosis. E2F-1 uses both p53-dependent and p53-independent pathways to kill cells. The p53-dependent pathway involves the induction by E2F-1 of the human tumour-suppressor protein p14ARF, which neutralizes HDM2 (human homologue of MDM2) and thereby stabilizes the p53 protein. Here we show that E2F-1 induces the transcription of the p53 homologue p73. Disruption of p73 function inhibited E2F-1-induced apoptosis in p53-defective tumour cells and in p53-/- mouse embryo fibroblasts. We conclude that activation of p73 provides a means for E2F-1 to induce death in the absence of p53.

Animals↗
PubMed2000

Human acid ceramidase is overexpressed but not mutated in prostate cancer.

The human acid ceramidase gene, that causes Farber disease, is located in 8p22, a region frequently altered in several cancers, including prostate cancer. Acid ceramidase catalyzes the hydrolysis of ceramide, a potent lipid second messenger molecule that promotes apoptosis and inhibits cellular proliferation. It is not known whether this gene, or its expression, is altered in prostate cancer. Here, we report the structural organization of the human gene, its expression in human tissues, and the identification of several single nucleotide polymorphisms. No cancer-related mutations were found in the gene in a panel of prostate tumor DNAs analyzed, but increased expression was observed in prostate tumor tissues when compared with matched normals. This increase was observed in all three prostate tumor cell lines tested (DU145, LnCAP, and PC3) when compared to a BPH (benign prostatic hyperplasia) cell line and 15/36 prostate tumors. These results suggest that acid ceramidase may play an important role in prostate carcinogenesis.

Acid Ceramidase↗
PubMed2000

Exon skipping in IVD RNA processing in isovaleric acidemia caused by point mutations in the coding region of the IVD gene.

Isovaleric acidemia (IVA) is a recessive disorder caused by a deficiency of isovaleryl-CoA dehydrogenase (IVD). We have reported elsewhere nine point mutations in the IVD gene in fibroblasts of patients with IVA, which lead to abnormalities in IVD protein processing and activity. In this report, we describe eight IVD gene mutations identified in seven IVA patients that result in abnormal splicing of IVD RNA. Four mutations in the coding region lead to aberrantly spliced mRNA species in patient fibroblasts. Three of these are amino acid altering point mutations, whereas one is a single-base insertion that leads to a shift in the reading frame of the mRNA. Two of the coding mutations strengthen pre-existing cryptic splice acceptors adjacent to the natural splice junctions and apparently interfere with exon recognition, resulting in exon skipping. This mechanism for missplicing has not been reported elsewhere. Four other mutations alter either the conserved gt or ag dinucleotide splice sites in the IVD gene. Exon skipping and cryptic splicing were confirmed by transfection of these mutations into a Cos-7 cell line model splicing system. Several of the mutations were predicted by individual information analysis to inactivate or significantly weaken adjacent donor or acceptor sites. The high frequency of splicing mutations identified in these patients is unusual, as is the finding of missplicing associated with missense mutations in exons. These results may lead to a better understanding of the phenotypic complexity of IVA, as well as provide insight into those factors important in defining intron/exon boundaries in vivo.

Animals↗
PubMed1998

Transcriptional regulation of mammalian cytochrome c oxidase genes.

The cytochrome c oxidase (COX) holoenzyme is a 13-subunit complex that carries out the terminal step in the electron transport chain. Three of the subunits, which contain the electron transfer function, are coded by mitochondrial DNA and the other ten subunits by nuclear DNA. Since the holoenzyme contains equivalent amounts of each subunit, we and others have examined transcriptional regulation of COX nuclear subunits to explore whether there is a common basis for co-regulation. Each gene is seen to have a unique pattern of recognition by regulatory factors; although some factors bind to more than one gene, not all COX genes seem to be regulated by the same set of factors. Current information about the COX promoters that have been examined is summarized, and the relation of promoter regulation to coordinate gene expression is discussed.

Animals↗
PubMed1997

Structural organization and promoter analysis of the bovine cytochrome c oxidase subunit VIIc gene. A functional role for YY1.

Cytochrome c oxidase (COX) subunit VIIc is one of the nuclear encoded subunits of the 13-subunit holoenzyme that carries out the terminal step in the electron transport chain. We have isolated the gene for this subunit, previously shown to be ubiquitously expressed from a single copy gene in the genome, and show that 167 base pairs of DNA surrounding the transcriptional start site contain the minimal promoter of this gene. This basal promoter contains two YY1 sites and at least one site for NRF-2, which show binding to their cognate factors. Mutation of both YY1 sites eliminates most of the promoter activity. Mutation at the upstream YY1 site significantly reduces the efficiency of transcript initiation at the major start site and thus plays the dominant role in COX7C regulation. COX7C is, thus, the second nuclear gene of COX that is regulated by YY1, suggesting that it is a third common factor, along with NRF-1 and NRF-2, to be associated with COX gene regulation.

Animals↗
PubMed1996

Cytochrome c oxidase subunit VIIa liver isoform. Characterization and identification of promoter elements in the bovine gene.

Cytochrome c oxidase subunit VIIa is specified by two nuclear genes, one (COX7AH) producing a heart/muscle-specific isoform and the other (COX7AL) a form expressed in all tissues. We have isolated both genes to examine their transcriptional regulation. Here, we characterize the core promoter of COX7AL and show that a 92-base pair region flanking the 5'-end promotes most of the activity of this gene. The 92-bp basal promoter contains sites for the nuclear respiratory factors NRF-1 and NRF-2, which have been shown to contribute to the transcription of a number of nuclear genes involved in mitochondrial respiratory activity, and also at least four Sp1 motifs. We show that both the NRF-1 and NRF-2 binding sites are functional in COX7AL and present evidence suggesting that interaction between the NRF-1 site and an upstream element contributes to expression.

Animals↗
PubMed1993

Structural organization and evolution of the liver isoform gene for bovine cytochrome c oxidase subunit VIIa.

Subunit VIIa of mammalian cytochrome c oxidase is one of three nuclear-encoded subunits that exhibit isoforms, existing predominantly as an H-form in cardiac and skeletal muscle tissues and as an L-form in others. We have isolated and characterized the L-isoform gene (COX7aL). It is 5.4 kb long, consists of four exons, and is located at a CpG island. Sp1 sites and an NRF1 site are located in an approximately 100-bp region immediately upstream of the gene. Comparison of the sequence and organization with the previously described H-isoform gene shows identical intron-exon organizations, with the first intron of both isoform genes splitting the presequence coding region almost identically. These results suggest that the isoform genes arose by duplication from a common ancestor prior to the mammalian radiation and that the ancestor already contained the presequences. In addition, four processed pseudogenes of the L-type have been isolated and characterized, one of which (COX7aLP1) contains no deletions, insertions, or frame-shifts and can encode a precursor protein of 83 amino acids. Construction of a phylogenetic tree employing extant COX7aL cDNA and bovine pseudogene sequences suggests that the expressed bovine gene and COX7aLP1 arose from a gene duplication event 4.6-6.8 Mya.

Amino Acid Sequence↗
PubMed1992

Tissue-specific expression and chromosome assignment of genes specifying two isoforms of subunit VIIa of human cytochrome c oxidase.

Subunit VIIa of mammalian cytochrome c oxidase (COX; EC 1.9.3.1) exists in at least two isoforms, one present in all tissue types ('liver' isoform; COX VIIa-L) and the other specific for cardiac and skeletal muscle (COX VIIa-M). We have isolated a full-length cDNA encoding human COX VIIa-M. The deduced polypeptide represents the human ortholog of COX VIIa-M, as it shares 78% identity with bovine COX VIIa-M, but only 63% identity with human COX VIIa-L. Northern-blot analysis of primate tissues demonstrated that COXVIIa-M mRNA is present only in muscle tissues; in contrast, the COXVIIa-L mRNA is present in both muscle and nonmuscle tissues. Southern-blot hybridization of human-rodent cell hybrid genomic DNA indicates that the COXVIIa-M gene maps to a single locus on chromosome 19, designated COX7AM. In contrast, COXVIIa-L cDNA probes hybridized to fragments from two COX7AL loci, on chromosomes 4 and 14.

Amino Acid Sequence↗
PubMed1992

Structure and organization of the heart isoform gene for bovine cytochrome c oxidase subunit VIIa.

Mammalian cytochrome c oxidase (COX) is a 13-subunit polypeptide complex that contains 10 subunits coded by the nucleus and 3 by the mitochondria. The nuclear-encoded subunits, though of unknown function, are presumed to play a regulatory role. Three of these (subunits VIa, VIIa, and VIII) generally exist in one of two isoforms--a constitutive (L) isoform or a skeletal muscle/heart-specific (H) isoform. To study the regulation, and possibly function, of these isoforms, we have begun characterizing the genes. In this paper we describe the isolation and characterization of the gene for the bovine COX VIIa-H isoform. The gene consists of four exons spanning 1.58 kb and is associated with a CpG island. There are no canonical TATA or CCAAT boxes immediately upstream of the transcription start site. Putative DNA sequence elements associated with respiratory function, muscle gene activation, and housekeeping function are present both in the upstream regions and within introns.

Amino Acid Sequence↗
PubMed1991

Cytochrome c oxidase subunit VIIa isoforms. Characterization and expression of bovine cDNAs.

Subunit VIIa of bovine cytochrome c oxidase occurs in two forms, the so-called heart and liver isoforms, which have been shown by protein analysis to differ in 35% of their amino acids. We have isolated and characterized cDNAs for each isoform. The derived heart-type processed protein is 59 amino acids long, with a 21-residue presequence; the immediate C terminus differs from the established protein sequence. The liver-type processed protein is 60 residues long, with a 23-amino acid presequence. Both presequences are traditional in that they are positively charged and appear amphiphilic when helically arrayed. The presequences are only 22% identical, but they both contain conserved residues indicative of two-step processing of the precursor proteins. Southern blot analysis reveals that the bovine genome contains five to six copies of the liver-type gene as opposed to the presence of a single copy heart-type gene. Transcriptional analysis shows that heart-type message is detectable only in heart and skeletal muscle; the liver type is also seen in heart and muscle and, additionally, in the other tissues examined (liver, brain, and lung). The amino acid sequence EKQKLFQED is conserved in rat and in both isoforms in cow and human and may represent a domain of core subunit function.

Amino Acid Sequence↗
PubMed1988

A processed pseudogene with an intact coding sequence for rat liver cytochrome c oxidase subunit VIc.

The biogenesis of eukaryotic cytochrome c oxidase involves the coordinate expression of nuclear and mitochondrial genes. Very little information is available on the gene structure of nuclear-coded cytochrome c oxidase subunits in mammalian systems. We report here the isolation and complete nucleotide sequence determination of a processed pseudogene for cytochrome c oxidase subunit VIc from rat liver. The pseudogene lacks introns and the coding region is intact with no deleterious lesions; however, there are 7 amino acid (aa) differences when compared to the sequence derived from cDNA clones. The pseudogene has the potential to code for a protein of 76 aa, containing a putative 3 aa N-terminal presequence when compared to the mature bovine heart VIc subunit. Potential regulatory regions, including a TATA box, are present in the 5'-flanking region.

Amino Acid Sequence↗
PubMed1986

Expression of cytochrome P-450 and albumin genes in rat liver: effect of xenobiotics.

Thioacetamide, a hepatocarcinogen and an inhibitor of heme synthesis, blocks the phenobarbitone-mediated increase in the transcription of cytochrome P-450b+e messenger RNA in rat liver. This property is also shared by CoCl2 and 3-amino-1,2,4-triazole, two other inhibitors of heme synthesis. Thus, it appears feasible that heme may serve as a positive regulator of cytochrome P-450b+e gene transcription. Thioacetamide enhances albumin messenger RNA concentration, whereas phenobarbitone decreases the same. However, these changes in albumin messenger RNA concentration are not accompanied by corresponding changes in the transcription rates. Therefore, drug-mediated changes in albumin messenger RNA concentration are due to posttranscriptional regulation. The property of thioacetamide to enhance the albumin messenger RNA concentration is not shared by CoCl2 and 3-amino-1,2,4-triazole. Therefore, heme does not appear to be a regulatory molecule mediating the reciprocal changes brought about in the concentrations of cytochrome P-450b+e and albumin messenger RNAs.

Acetamides↗
PubMed1984

Construction of a cDNA clone for a nuclear-coded subunit of cytochrome c oxidase from rat liver.

A cDNA library for 6S-9S poly(A)-containing RNA from rat liver was constructed in E. coli. Initial screening of the clones was carried out using single stranded 32P-labeled cDNA prepared against poly(A)-containing RNA isolated from immunoadsorbed polyribosomes enriched for the nuclear-coded subunit messenger RNAs of cytochrome c oxidase. One of the clones, pCO89, was found to hybridize with the messenger RNA for subunit VIC. The DNA sequence of the insert in pCO89 was carried out and it has got extensive homology with the C-terminal 33 amino acids of subunit VIC from beef heart cytochrome c oxidase. In addition, the insert contained 146 bp, corresponding to a portion of the 3'-non-coding region. Northern blot analysis of rat liver RNA with the nick-translated insert of pCO89 revealed that the messenger RNA for subunit VI would contain around 510 bases.

Animals↗