Human molybdopterin synthase gene: identification of a bicistronic transcript with overlapping reading frames. (1/234)

A universal molybdenum-containing cofactor (MoCo) is essential for the activity of all human molybdoenzymes, including sulphite oxidase. The free cofactor is highly unstable, and all organisms share a similar biosynthetic pathway. The involved enzymes exhibit homologies, even between bacteria and humans. We have exploited these homologies to isolate a cDNA for the heterodimeric molybdopterin (MPT)-synthase. This enzyme is necessary for the conversion of an unstable precursor into molybdopterin, the organic moiety of MoCo. The corresponding transcript shows a bicistronic structure, encoding the small and large subunits of the MPT-synthase in two different open reading frames (ORFs) that overlap by 77 nucleotides. In various human tissues, only one size of mRNA coinciding with the bicistronic transcript was detected. In vitro translation and mutagenesis experiments demonstrated that each ORF is translated independently, leading to the synthesis of a 10-kDa protein and a 21-kDa protein for the small and large subunits, respectively, and indicated that the 3'-proximal ORF of the bicistronic transcript is translated by leaky scanning.  (+info)

Tbx5 is essential for heart development. (2/234)

Mutations in the Tbx5 transcription factor cause heart septal defects found in human Holt-Oram Syndrome. The complete extent to which Tbx5 functions in heart development, however, has not been established. Here we show that, in Xenopus embryos, Tbx5 is expressed in the early heart field, posterior to the cardiac homeobox transcription factor, Nkx2.5. During morphogenesis, Tbx5 is expressed throughout the heart tube except the anterior portion, the bulbus cordis. When Tbx5 activity is antagonized with a hormone-inducible, dominant negative version of the protein, the heart fails to develop. These results suggest that, in addition to its function in heart septation, Tbx5 has a more global role in cardiac specification and heart development in vertebrate embryos.  (+info)

Comparative study of overlapping genes in the genomes of Mycoplasma genitalium and Mycoplasma pneumoniae. (3/234)

Overlapping genes are defined, in this paper, as a pair of adjacent genes whose coding regions are partly overlapping. We systematically analyzed all overlapping genes in the genomes of two closely related species: Mycoplasma genitalium and Mycoplasma pneumoniae. Careful comparisons were made for homologous genes that are overlapped in one species but not in the other. This comparative analysis allows us to propose a model of how overlapping genes emerged in the course of evolution. It was found that overlapping genes were generated primarily due to the loss of a stop codon in either gene, in many cases, the absence of which resulted in elongation of the 3' end of the gene's coding region. More specifically, the loss of the stop codon took place as a result of the following events: deletion of the stop codon (64.4%), point mutation at the stop codon (4.4%), and frame shift at the end of the coding region (6.7%). Overlapping genes, in a sense, can be thought of as the results of evolutionary pressure to minimize genome size. However, our analysis indicates that many overlapping genes, at least in the genomes of M.genitalium and M.pneumoniae, are due to incidental elongation of the coding regions.  (+info)

Alternate choice of initiation codon produces a biologically active product of the von Hippel Lindau gene with tumor suppressor activity. (4/234)

The VHL tumor suppressor gene has previously been reported to encode a protein of 213 amino acid residues. Here we report the identification of a second major VHL gene product with an apparent molecular weight of 18 kD, pVHL18, which appears to arise from alternate translation initiation at a second AUG codon (codon 54) within the VHL open reading frame. In vitro and in vivo studies indicate that the internal codon in the VHL mRNA is necessary and sufficient for production of pVHL18. pVHL18 can bind to elongin B, elongin C, and Hs-CUL2. When reintroduced into renal carcinoma cells that lack a wild-type VHL allele, pVHL18 suppresses basal levels of VEGF expression, restores hypoxia-inducibility of VEGF expression, and inhibits tumor formation in nude mice. These data strongly support the existence of two distinct VHL gene products in VHL tumor suppression.  (+info)

Identification of a surface of FNR overlapping activating region 1 that is required for repression of gene expression. (5/234)

A library of Escherichia coli fnr mutants has been screened to identify FNR (regulator of fumarate and nitrate reduction) variants that are defective repressors, but competent activators. All but one of seventeen variants had substitutions close to or within the face of FNR that contains activating region 1 (AR1). Activating region 1 is known to contact the alpha subunit of RNA polymerase to facilitate transcription activation. It is now evident that this face also has a role in FNR-mediated repression. Single amino acid substitutions at Lys54, Gly74, Ala95, Met147, Leu193, Arg197, or Leu239, and double substitutions at Ser13 and Ser145, Cys16 and Ile45, Tyr69 and Ser133, or Lys164 and Phe191, impaired FNR-mediated repression of ndh without greatly affecting activation from model Class I (FNR site at -71.5) and Class II (FNR site at -41.5) FNR-activated promoters. Although repression was impaired in a second group of FNR variants with substitutions at Leu34, Arg72 and Leu193, Phe92, or Ser178, transcription activation from the simple FNR-dependent promoters was severely reduced. However, expression from pyfiD (FNR sites at -40.5 and -93.5) and a derivative lacking the site at -93.5, pyfiD-/+, remained relatively high indicating that this second group have a context-dependent activation defect as well as a repression defect. The prediction that the substitutions affecting repression were likely to be in solvent exposed regions of FNR was supported by analysis of peptides produced by partial proteolysis of FNR. Thus, FNR-mediated repression at promoters with multiple FNR sites requires regions of FNR that are different from, but overlap, AR1.  (+info)

Functional anatomy of herpes simplex virus 1 overlapping genes encoding infected-cell protein 22 and US1.5 protein. (6/234)

Earlier studies have shown that (i) the coding domain of the alpha22 gene encodes two proteins, the 420-amino-acid infected-cell protein 22 (ICP22) and a protein, US1.5, which is initiated from methionine 147 of ICP22 and which is colinear with the remaining portion of that protein; (ii) posttranslational processing of ICP22 mediated largely by the viral protein kinase UL13 yields several isoforms differing in electrophoretic mobility; and (iii) mutants lacking the carboxyl-terminal half of the ICP22 and therefore DeltaUS1.5 are avirulent and fail to express normal levels of subsets of both alpha (e.g., ICP0) or gamma2 (e.g., US11 and UL38) proteins. We have generated and analyzed two sets of recombinant viruses. The first lacked portions of or all of the sequences expressed solely by ICP22. The second set lacked 10 to 40 3'-terminal codons of ICP22 and US1. 5. The results were as follows. (i) In cells infected with mutants lacking amino-terminal sequences, translation initiation begins at methionine 147. The resulting protein cannot be differentiated in mobility from authentic US1.5, and its posttranslational processing is mediated by the UL13 protein kinase. (ii) Expression of US11 and UL38 genes by mutants carrying only the US1.5 gene is similar to that of wild-type parent virus. (iii) Mutants which express only US1. 5 protein are avirulent in mice. (iv) The coding sequences Met147 to Met171 are essential for posttranslational processing of the US1.5 protein. (v) ICP22 made by mutants lacking 15 or fewer of the 3'-terminal codons are posttranslationally processed whereas those lacking 18 or more codons are not processed. (vi) Wild-type and mutant ICP22 proteins localized in both nucleus and cytoplasm irrespective of posttranslational processing. We conclude that ICP22 encodes two sets of functions, one in the amino terminus unique to ICP22 and one shared by ICP22 and US1.5. These functions are required for viral replication in experimental animals. US1.5 protein must be posttranslationally modified by the UL13 protein kinase to enable expression of a subset of late genes exemplified by UL38 and US11. Posttranslational processing is determined by two sets of sequences, at the amino terminus and at the carboxyl terminus of US1.5, respectively, a finding consistent with the hypothesis that both domains interact with protein partners for specific functions.  (+info)

A novel imprinted gene, encoding a RING zinc-finger protein, and overlapping antisense transcript in the Prader-Willi syndrome critical region. (7/234)

We describe a complex imprinted locus in chromosome 15q11-q13 that encodes two genes, ZNF127 and ZNF127AS. The ZNF127 gene encodes a protein with a RING (C3HC4) zinc-finger and multiple C3H zinc-finger motifs, the former being closely related to a protein from variola major virus, the smallpox etiological agent. These motifs allow prediction of ZNF127 function as a ribonucleoprotein. The intronless ZNF127 gene is expressed ubiquitously, but the entire coding sequence and 5' CpG island overlaps a second gene, ZNF127AS, that is transcribed from the antisense strand with a different transcript size and pattern of expression. Allele-specific analysis shows that ZNF127 is expressed only from the paternal allele. Consistent with this expression pattern, in the brain the ZNF127 5' CpG island is completely unmethylated on the paternal allele but methylated on the maternal allele. Analyses of adult testis, sperm and fetal oocytes demonstrates a gametic methylation imprint with unmethylated paternal germ cells. Recent findings indicate that ZNF127 is part of the coordinately regulated imprinted domain affected in Prader-Willi syndrome patients with imprinting mutations. Therefore, ZNF127 and ZNF127AS are novel imprinted genes that may be associated with some of the clinical features of the polygenic Prader-Willi syndrome.  (+info)

Origin of gene overlap: the case of TCP1 and ACAT2. (8/234)

The human acetyl-CoA acetyltransferase 2 gene, ACAT2, codes for a thiolase, an enzyme involved in lipid metabolism. The human T-complex protein 1 gene, TCP1, encodes a molecular chaperone of the chaperonin family. The two genes overlap by their 3'-untranslated regions, their coding sequences being located on opposite DNA strands in a tail-to-tail orientation. To find out how the overlap might have arisen in evolution, the homologous genes of the zebrafish, the African toad, caiman, platypus, opossum, and wallaby were identified. In each species, standard or long polymerase chain reactions were used to determine whether the ACAT2 and TCP1 homologs are closely linked and, if so, whether they overlap. The results reveal that the overlap apparently arose during the transition from therapsid reptiles to mammals and has been retained for >200 million years. Part of the overlapping untranslated region shows remarkable sequence conservation. The overlap presumably arose during the chromosomal rearrangement that brought the two unrelated and previously separated genes together. One or both of the transposed genes found by chance signals that are necessary for the processing of their transcripts to be present on the noncoding strand of the partner gene.  (+info)

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