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PubMed1980

Overlapping genes at the cheA locus of Escherichia coli.

The cheA locus of Escherichia coli, which is required for chemotactic behavior, encodes two polypeptide products designated p[cheA]L and p[cheA]S. The mode of synthesis of these two proteins was investigated by transferring various missense and nonsense mutations to a lambda transducing phage and observing the mutant cheA products made after infection of ultraviolet-irradiated host cells. Missense mutations had no effect on either the size or the relative amounts of the two cheA polypeptides. Most nonsense mutations caused premature translational termination of both cheA products, indicating that p[cheA]L and p[cheA]S must be translated from the same coding sequence in the same reading frame. Two exceptional nonsense alleles at the promoter-proximal end of cheA made an intact p[cheA]s but no detectable p[cheA]L. These findings show that the cheA locus may contain two different sites for initiation of translation. The synthesis of both proteins can be effected by the same promoter, but it is not yet clear whether both are translated from identical mRNA molecules. Complementation studies of cheA mutants provided evidence for two functional activities, one associated with the amino terminus of p[cheA]L and the other with the common portions of p[cheA]L and p[cheA]S. It is possible that each cheA product has a different function required for chemotaxis. The possible roles of these two products and the functional significance of bacterial genes with overlapping coding sequences are discussed.

Alleles↗
PubMed2003

Peas-Mea1-Ppp2r5d overlapping gene complex: a transposon mediated-gene formation in mammals.

Human and mouse MEA1/Mea1 is flanked by two overlapping genes, a novel PEAS/Peas in a head-to-head orientation and PPP2R5D/Ppp2r5d in a tail-to-tail orientation making a Peas-Mea1-Ppp2r5d overlapping gene complex (PMP-complex). Genomic zoo blot analyses and database searching revealed that Mea1 exists only in mammals, while Peas and Ppp2r5d are conserved in eukaryotes. Mea1 and Peas are transcribed from a testis-expressed bidirectional promoter. Mea1-Ppp2r5d overlapping segment (MPOS) contains polyadenylation signals for both genes and shows marked conservation throughout mammals. Furthermore, the MPOS occupies 3'-region of transcripts of both genes is expected to form a clover-like intramolecular structure. Mouse genomic library Screening and database searches identified two MPOS-derived sequences in Odf2 gene and RP23-86H7 cosmid clone, respectively, in which MPOS might be a core segment for the retropositions. Thus, a key role of MPOS, a short transposable element containing polyadenylation signals on both strands, in the formation of the Mea1 during mammalian evolution is suggested.

Animals↗
PubMed2006

A CTCF-dependent silencer located in the differentially methylated area may regulate expression of a housekeeping gene overlapping a tissue-specific gene domain.

The tissue-specific chicken alpha-globin gene domain represents one of the paradigms, in terms of its constitutively open chromatin conformation and the location of several regulatory elements within the neighboring housekeeping gene. Here, we show that an 0.2-kb DNA fragment located approximately 4 kb upstream to the chicken alpha-globin gene cluster contains a binding site for the multifunctional protein factor CTCF and possesses silencer activity which depends on CTCF binding, as demonstrated by site-directed mutagenesis of the CTCF recognition sequence. CTCF was found to be associated with this recognition site in erythroid cells but not in lymphoid cells where the site is methylated. A functional promoter directing the transcription of the apparently housekeeping ggPRX gene was found 120 bp from the CTCF-dependent silencer. The data are discussed in terms of the hypothesis that the CTCF-dependent silencer stabilizes the level of ggPRX gene transcription in erythroid cells where the promoter of this gene may be influenced by positive cis-regulatory signals activating alpha-globin gene transcription.

Animals↗
PubMed1987

Gene overlap and site-specific attenuation of transcription of the viral polymerase L gene of human respiratory syncytial virus.

Sequence analysis of the gene encoding polymerase L protein of respiratory syncytial virus showed that L-gene transcription initiates within its upstream neighbor, the gene encoding the 22-kDa protein 22K. This is an exception to the canonical mode of sequential transcription of nonoverlapping genes described for other nonsegmented negative-strand RNA viruses. As a consequence of the gene overlap, the termination/polyadenylylation signal for the 22K gene is located at nucleotides 56-68 within the L gene. L-gene transcription yielded two RNAs: an abundant, truncated, polyadenylylated transcript resulting from termination at the internal signal and a markedly less abundant large polyadenylylated transcript representing the complete L gene. This result showed that the internal termination/polyadenylylation signal is an attenuator of L-gene transcription.

Amino Acid Sequence↗
PubMed2004

The light-responsive promoter of cyanobacterial ORF76 gene overlaps with the htpG terminator.

A Synechococcus PCC7942 gene, encoding 76 amino acids of an unknown protein (designated ORF76), is located at the same orientation and downstream of the htpG gene. The processed site of ORF76 transcripts is located at position +39 with respect to the true transcription initiation site. The non-E. colisigma70-like basal promoter of ORF76 (-51 to -9) is controlled by three cis-acting elements: positively acting element (-160 to -86), negative regulatory element (-86 to -51) and light-responsive element (-51 to +63) that, together, respond to high light induction at transcriptional level. The ORF76 gene is expressed as monocistronic transcript. The promoter elements of ORF76 gene overlap with the coding sequence and 3' end formation signal of htpG gene.

Amino Acid Sequence↗
PubMed2005

Overlapping gene mutations of hepatitis B virus in a chronic hepatitis B patient with hepatitis B surface antigen loss during lamivudine therapy.

Disappearance of hepatitis B surface antigens (HBsAg) in chronic hepatitis B usually indicates clearance of hepatitis B virus (HBV) infection. However, false HBsAg negativity with mutations in pre-S2 and 'a' determinant has been reported. It is also known that YMDD mutations decrease the production of HBV and escape detection of serum HBsAg. Here, we report overlapping gene mutations in a patient with HBsAg loss during the lamivudine therapy. After 36 months of lamivudine therapy in a 44-yrold Korean chronic hepatitis B patient, serum HBsAg turned negative while HBV DNA remained positive by a DNA probe method. Nucleotide sequence of serum HBV DNA was compared with the HBV genotype C subtype adr registered in NCBI AF 286594. Deletion of nucleotides 23 to 55 (amino acids 12 to 22) was identified in the pre-S2 region. Sequencing of the 'a' determinant revealed amino acid substitutions as I126S, T131N, M133T, and S136Y. Methionine of rtM204 in the P gene was substituted for isoleucine indicating YIDD mutation (rtM204I). We identified a HBV mutant composed of pre-S2 deletions and 'a' determinant substitutions with YMDD mutation. Our result suggests that false HBsAg negativity can be induced by combination of overlapping gene mutations during the lamivudine therapy.

Adult↗
PubMed2004

FISHing for chick genes: Triple-label whole-mount fluorescence in situ hybridization detects simultaneous and overlapping gene expression in avian embryos.

Multi-color whole-mount in situ hybridization is a powerful technique for comparing the spatial expression patterns of two or more genes in developing embryos. We have developed an amplified triple-label whole-mount fluorescence in situ hybridization (FISH) protocol that permits detection of three different mRNAs in a single embryo. Our protocol uses simultaneous in situ hybridization to haptenylated riboprobes, followed by sequential antibody detection using anti-hapten antibodies conjugated to horseradish peroxidase, and the tyramide signal amplification (TSA) fluorescence detection system. Conventional fluorescence microscopy identifies areas of overlapping gene expression at the tissue level, whereas confocal fluorescence microscopy permits single-cell resolution and differentiates specialized cell types within a given tissue. This protocol will provide researchers engaged in the use of FISH with a solid starting point for adapting their own in situ hybridization protocols, either alone or in combination with immunohistochemistry or green fluorescence protein colocalization.

Animals↗
PubMed1999

Origin of gene overlap: the case of TCP1 and ACAT2.

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.

Acetyl-CoA C-Acetyltransferase↗
PubMed1984

Gene X of bacteriophage f1 is required for phage DNA synthesis. Mutagenesis of in-frame overlapping genes.

The gene II protein of bacteriophage f1 is a site-specific endonuclease required for initiation of phage viral strand DNA synthesis. Within gene II is another gene, X, encoding a protein of unknown function identical to the C-terminal 27% of the gene II protein, and separately translated from codon 300 (AUG) of gene II. By oligonucleotide mutagenesis, we constructed phage mutants in which this codon has been changed to UAG (amber) or UUG (leucine), and propagated them on cells carrying a cloned copy of gene X on a plasmid. The amber mutant makes no gene X protein, and cannot grow in the absence of the complementing plasmid; the leucine-inserting mutant can make gene X protein, and grows normally without the plasmid. Without gene X protein, phage DNA synthesis (particularly viral strand synthesis) is impaired. We discuss this finding in the context of other known in-frame overlapping genes (particularly genes A and A* of phage phi X174), many of which are also involved in the specific initiation of DNA synthesis, and suggest applications for the mutagenic strategy we employed.

Coliphages↗
PubMed2005

A high frequency of overlapping gene expression in compacted eukaryotic genomes.

The gene density of eukaryotic nuclear genomes is generally low relative to prokaryotes, but several eukaryotic lineages (many parasites or endosymbionts) have independently evolved highly compacted, gene-dense genomes. The best studied of these are the microsporidia, highly adapted fungal parasites, and the nucleomorphs, relict nuclei of endosymbiotic algae found in cryptomonads and chlorarachniophytes. These systems are now models for the effects of compaction on the form and dynamics of the nuclear genome. Here we report a large-scale investigation of gene expression from compacted eukaryotic genomes. We have conducted EST surveys of the microsporidian Antonospora locustae and nucleomorphs of the cryptomonad Guillardia theta and the chlorarachniophyte Bigelowiella natans. In all three systems we find a high frequency of mRNA molecules that encode sequence from more than one gene. There is no bias for these genes to be on the same strand, so it is unlikely that these mRNAs represent operons. Instead, compaction appears to have reduced the intergenic regions to such an extent that control elements like promoters and terminators have been forced into or beyond adjacent genes, resulting in long untranslated regions that encode other genes. Normally, transcriptional overlap can interfere with expression of a gene, but these genomes cope with high frequencies of overlap and with termination signals within expressed genes. These findings also point to serious practical difficulties in studying expression in compacted genomes, because many techniques, such as arrays or serial analysis of gene expression will be misleading.

Animals↗
PubMed2006

Overlapping genes as rare genomic markers: the phylogeny of gamma-Proteobacteria as a case study.

Phylogenies can be constructed in many ways, including using shared complex characters known as rare genomic changes (RGCs), such as insertions and deletions (indels), retroposon integrations and intron positions. Here, we demonstrate that distance-based phylogenies, which were determined by shared overlapping genes from 13 completely sequenced gamma-Proteobacteria genomes, are consistent with phylogenies based on 16S rRNAs and other robust markers. These findings suggest that overlapping genes could provide interesting additional insights into the phylogenomics of completely sequenced microbial genomes.

Algorithms↗
PubMedDate not supplied

[Creation of artificial hybrid operons with partially overlapping genes to achieve an expression of heterologous genes in Escherichia coli cells].

A new method of optimization of foreign gene expression in E. coli, based on the construction of hybrid operons with partially overlapping genes is described. The partial overlapping of the translation termination and initiation sites in the formed operon must provide translational coupling of appropriate gene product synthesis. Such an approach has provided the synthesis of human interferon alpha F in E. coli cells under the control of the lacUV5-promotor up to about (3-4).10(7) units per liter of bacterial culture. The reinitiation of the distal gene translation is shown to take place in the intercistronic region. Substitution of the lacUV5 promotor by the more efficient tac one allowed to increase the synthesis level of interferon alpha F to (1-2).10(8) units per liter. The conclusion is made about the equimolarity of distal and proximal to the promotor genes products syntheses when the intercistronic region of E. coli trpE-trpD genes are used for translational coupling.

Base Sequence↗
PubMed1988

Mechanism of expression of the overlapping genes of Bacillus subtilis aspartokinase II.

The mechanism of expression of the overlapping genes that encode the alpha and beta subunits of aspartokinase II of Bacillus subtilis was studied by specific mutagenesis of the cloned coding sequence. Escherichia coli or B. subtilis VB31 (aspartokinase II-deficient), transformed with plasmids carrying either a deletion of the translation start site and about one-half of the coding region for the larger alpha subunit or a frameshift mutation early in the alpha subunit coding region, produced the smaller beta subunit in the absence of alpha subunit synthesis, indicating that beta subunit is not derived from alpha subunit and that its synthesis does not depend on the alpha subunit translation initiation site. The beta subunit translation start site was identified by oligonucleotide-directed mutagenesis of the putative translation start codon. Modification of the nucleotide sequence encoding methionine residue 247 of the alpha subunit from ATG to either TTA or AAT (but not GTG) abolished beta subunit synthesis but had no effect on the production of alpha subunit. This observation is consistent with peptide chain initiation by N-formylmethionine, which specifically requires an ATG or GTG sequence, and indicates that translation of the beta subunit starts at a site corresponding to Met247 of the alpha subunit. Initial studies on the function of the aspartokinase II subunits, using E. coli as a heterologous host, showed that beta subunit was not essential for the expression of the catalytic function of aspartokinase, measured in vitro and in vivo, nor for its allosteric regulation by L-lysine. Whether the beta subunit has a function specific to B. subtilis needs to be explored in a homologous expression system.

Aspartate Kinase↗
PubMed1988

The A and B subgroups of human respiratory syncytial virus: comparison of intergenic and gene-overlap sequences.

Intergenic and flanking gene regions for the 1C-1B, 1B-N, N-P, M-1A, G-F and F-22K gene junctions of respiratory syncytial virus strain 18537, representing antigenic subgroup B, were determined by dideoxynucleotide sequencing of polycistronic cDNAs and mRNAs. Comparison with their counterparts from the subgroup A strain A2 showed that the intergenic sequences were not conserved within or between the strains. Flanking non-coding gene sequences also were generally not conserved except for the highly conserved gene-start and gene-end transcription signals. The sequence of the overlap between the 22K and L genes was conserved almost exactly between the two subgroups.

Base Sequence↗
PubMed1997

Constrained evolution with respect to gene overlap of hepatitis B virus.

With the aim of elucidating the evolution of a hepadnavirus family, we constructed molecular phylogenetic trees for 27 strains of hepatitis B virus (HBV) using both the unweighted pair-grouping and neighbor-joining methods. All five gene regions, P, C, S, X, and preS, were used to construct the phylogenetic trees. Using the phylogenetic trees obtained, we classified these strains into five major groups in which the strains were closely related to each other. Our classification reinforced our previous view that genetic classification is not always compatible with conventional classification determined by serological subtypes. Moreover, constraints on the evolutionary process of HBV were analyzed for amino-acid-altering (nonsynonymous) and silent (synonymous) substitutions, because two-thirds of the open reading frame (ORF), P, contains alternating overlapping ORFs. In our unique analysis of this interesting gene structure of HBV, the most frequent synonymous substitutions were observed in the nonoverlapped parts of the P and C genes. On the other hand, the number of synonymous substitutions per nucleotide site for the S gene was quite low and appeared a strongly constrained evolution. Because the P gene overlaps the S gene in a different frame, the low rate of synonymous substitution for the S gene can be explained by the evolutionary constraints which are imposed on the overlapping gene region. In other words, synonymous substitutions in the S gene can cause amino acid changes in its overlapping region in a different frame. Thus, the evolution of HBV is constrained evolutionarily by the overlapping genes. We propose calling this mode of viral evolution "constrained evolution." The evolution of HBV represents a typical constrained evolution.

Amino Acid Sequence↗