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PubMed1994

Identification of genes encoding ribosomal protein L33 from Bacillus licheniformis, Thermus thermophilus and Thermotoga maritima.

Three previously unrecognized genes from Bacillus licheniformis, Thermus thermophilus and Thermotoga maritima, encoding ribosomal protein L33, have been identified and designated as rpmG genes. Their sequence and context have been compared with the three known rpmG genes from Escherichia coli, Lactococcus lactis and B. subtilis. Previously unrecognized open reading frames homologous to secE are located immediately downstream from rpmG in B. licheniformis, T. thermophilus and Ta. maritima: beyond that lie previously identified nusG and rplK genes. Thus, the genes located downstream from rpmG show similar organization across three of the four bacterial phyla represented.

Amino Acid Sequence↗

PubMed1995

Cloning and sequencing of a gene from the archaeon Pyrococcus furiosus with high homology to a gene encoding phosphoenolpyruvate synthetase from Escherichia coli.

A gene from the hyperthermophilic archaeon Pyrococcus furiosus, strain Vc1 (DSM 3638), contains an 817-amino-acid open reading frame which shows 42% identity to the phosphoenolpyruvate (PEP) synthetase of Escherichia coli. This putative P. furiosus PEP synthetase is slightly larger than the E. coli enzyme, the region between residues 58 and 89 being absent from the latter.

Amino Acid Sequence↗

PubMed2005

uAUG and uORFs in human and rodent 5'untranslated mRNAs.

The control of translation is a fundamental mechanism in the regulation of gene expression. Among the cis-acting elements that play a role in translation regulation are upstream open reading frames (uORFs) and upstream AUG (uAUGs) located in the 5'UTR of mRNAs. We present here a genome-wide analysis of uAUGs and uORFs in a curated set of human and rodent mRNAs. Our study shows that the occurrence of uAUGs is suppressed more strongly than that of uORFs and that in-frame uAUGs are more strongly suppressed than out-of-frame uAUGs. A very similar pattern of uAUG/uORF frequency was also observed in mouse mRNAs. The analysis of orthologous 5'UTR sequences revealed a remarkable degree of evolutionary conservation only of those uORFs which acquired some functional activity. Our data suggest that besides leaky scanning and reinitiation, which likely occur with variable and gene-specific efficiency, the ribosome-shunt mechanism, eventually coupled to reinitiation after uORF translation, may be a widespread mode of translation regulation in eukaryotes.

5' Untranslated Regions↗

PubMed2004

Identification of BRCA1-IRIS, a BRCA1 locus product.

Breast cancer is the most common malignancy among women, and mutations in the BRCA genes produce increased susceptibility to these malignancies in certain families. Here we identify BRCA1-IRIS as a 1,399-amino-acid BRCA1 gene product encoded by an uninterrupted open reading frame that extends from codon 1 of the known BRCA1 open reading frame to a termination point 34 triplets into intron 11. Unlike full-length BRCA1 (p220), BRCA1-IRIS is exclusively chromatin-associated, fails to interact with BARD1 in vivo or in vitro and exhibits unique nuclear immunostaining. Unlike BRCA1FL (or p220), BRCA1-IRIS also co-immunoprecipitated with DNA-replication-licensing proteins and with known replication initiation sites. Suppression of BRCA1-IRIS expression hindered the normal departure of geminin from pre-replication complexes, and depressed the rate of cellular DNA replication and possibly initiation-related synthesis. In contrast, BRCA1-IRIS overexpression stimulated DNA replication. These data imply that endogenous BRCA1-IRIS positively influences the DNA replication initiation machinery.

Amino Acid Sequence↗

PubMed1992

Characterization of the nucleotide sequence of the Lymantria dispar nuclear polyhedrosis virus DNA polymerase gene region.

The DNA polymerase gene of the Lymantria dispar multinucleocapsid nuclear polyhedrosis virus (LdMNPV) was cloned and sequenced. The predicted DNA polymerase protein (1113 amino acids, 115.9K) was found to have an amino acid identity of 48% with the corresponding gene of the Autographa californica MNPV (AcMNPV). It contains five domains associated with substrate binding, primase interaction, and pyrophosphate hydrolysis and three domains associated with 3'-->5' exonuclease activity common to other DNA polymerases. A region with a conserved TATA promoter and a CAGT mRNA start site sequence motif was identified and shown to be transcribed by RNA polymerase II, indicating that the LdMNPV DNA polymerase gene is expressed as an early gene. An open reading frame possibly expressed as a late gene, oriented in the opposite direction and overlapping the N-terminal coding region of the DNA polymerase gene was found in the LdMNPV sequence and was shown to be conserved in the same position in AcMNPV.

Amino Acid Sequence↗

PubMed2005

Post-transcriptional control of a glucoamylase gene from Trichoderma harzianum under stress conditions.

We have examined the regulation of Trichoderma harzianum glucoamylase gene (Gla66) in response to different growth conditions. Transcription of the Gla66 gene is initiated from two different sites, yielding two transcripts of 2.1 kb and 2.6 kb respectively. The 2.1 kb mRNA (ST) encodes for an extracellular glucoamylase of 66 kDa. This protein shows the domains conserved in other fungal glucoamylases: a signal peptide responsible for protein secretion and a catalytic domain, both joined by a linker region. The longest transcript (LT) is untranslated, it contains an unusually extended 5'-untranslated region and is transcribed under stress and growth limiting conditions. The translational control of LT could be defined by the presence of four upstream open reading frames (uORFs) in its 5'-leader sequence. The analysis of these uORFs in a yeast heterologous system shows that two of these uORFs prevent the Gla66 translation under unfavourable growth conditions, when the LT transcript is accumulated.

5' Untranslated Regions↗

PubMed2000

Genetic characterization of vanG, a novel vancomycin resistance locus of Enterococcus faecalis.

Enterococcus faecalis strain WCH9 displays a moderate level of resistance to vancomycin (MIC = 16 microgram/ml) and full susceptibility to teicoplanin but is negative by PCR analysis using primers specific for all known enterococcal vancomycin resistance genotypes (vanA, vanB, vanC, vanD, and vanE). We have isolated and sequenced a novel putative vancomycin resistance locus (designated vanG), which contains seven open reading frames, from this strain. These are organized differently from those of all the other enterococcal van loci, and, furthermore, the individual vanG gene products exhibit less than 50% amino acid sequence identity to other van gene products.

Amino Acid Sequence↗

PubMed2004

Molecular characterization of tgd057, a novel gene from Toxoplasma gondii.

The expressed sequence tag (EST) effort in Toxoplasma gondii has generated a substantial amount of gene information. To exploit this valuable resource, we chose to study tgd057, a novel gene identified by a large number of ESTs that otherwise show no significant match to known sequences in the database. Northern analysis showed that tgd057 is transcribed in this tachyzoite. The complete cDNA sequence of tgd057 is 1169 bp in length. Sequence analysis revealed that tgd057 possibly adopts two polyadenylation sites, utilizes the fourth in-frame ATG for translation initiation, and codes for a secretory protein. The longest open reading frame for the tgd057 gene was cloned and expressed as a recombinant protein (rd57) in Escherichia coli. Western analysis revealed that serum against rd57 recognized a molecule of ~21 kDa in the tachyzoite protein extract. This suggests that the tgd057 gene is expressed in vivo in the parasite.

Amino Acid Sequence↗

PubMed2006

Cloning, chromosomal characterization and mapping of the NAD-dependent histone deacetylases gene sirtuin 1.

Sirtuin 1 (SIRT1) is a nicotinamide adenine dinucleotide (NAD+)-dependent deacetylase, which belongs to the silent information regulator 2 (Sir2) family of sirtuin histone deacetylases (HDACs). The yeast Sir2 protein and its mammalian derivatives play a central role in epigenetic gene silencing, DNA repair and recombination, cell-cycle, microtubule organization, and in the regulation of aging. We have isolated and characterized the human Sirt1 genomic sequence, which spans a region of 33,660 bp and which has one single genomic locus. Determination of the exon-intron splice junctions established that SIRT1 is encoded by 9 exons ranging in size from 80 bp (exon 6) to 2,120 bp (exon 9). Characterization of the 5' flanking genomic region, which precedes the Sirt1 open reading frame, revealed a CCAAT-box and a number of NF-kappaB and GATA transcription factor binding sites in addition to a small 350 bp CpG island. The 4,107 bp human Sirt1 mRNA has an open reading frame of 2,244 bp and encodes a 747 aa protein with a predictive molecular weight of 81.7 kDa and an isoelectric point of 4.55. Fluorescence in situ hybridization analysis localized the human Sirt1 gene to chromosome 10q21.3.

Amino Acid Sequence↗

PubMed1990

[Confirmation of the presence of a transcription termination site in a DNA segment downstream from gene 31 of bacteriophage T4].

Two open reading frames followed by a rho-independent transcription terminator and early promoter were established in a sequenced region downstream from gene 31 of T4 phage. The mRNA hairpin suggested by the terminator sequence has the structure (x)8CUUCGG(y)8, the loop sequence UUCG having been shown to significantly stabilize RNA hairpins.

Amino Acid Sequence↗

PubMed1996

Sea urchin genes expressed in activated coelomocytes are identified by expressed sequence tags. Complement homologues and other putative immune response genes suggest immune system homology within the deuterostomes.

To identify some of the genes expressed in LPS-activated coelomocytes, we sequenced randomly chosen clones from a directionally constructed cDNA library to produce a set of expressed sequence tags (ESTs). Deduced amino acid sequences from 307 ESTs were compared with known protein sequences in GenBank, and significant matches to approximately 30% of the clones were identified. Eighty-nine clones matched to 55 different proteins, including several putative immune effector proteins. In this work, we show the first identification of an invertebrate homologue of a vertebrate C component. Another EST matches to several short consensus repeats that are characteristic of a variety of proteins, including CR/regulatory proteins and clotting factors. Additional putative immune effector genes include 1) a Kazal-type protease inhibitor that may function to inactivate bacterial proteases, 2) a C-type lectin similar to echinoidin, and 3) a serine protease with similarities to thrombin, elastase, haptoglobin, and plasmin. Other EST categories include 1) cell surface proteins and receptors, 2) proteins involved in signaling systems, 3) lysosomal and secreted proteins, 4) cytoskeletal and cytoskeletal modifying proteins, 5) general cell function proteins, 6) proteins with unknown function, and 7) ESTs without significant matches, 25 with open reading frames. Many of the ESTs identified in this study represent the types of genes expected to be used in lower deuterostome immune functions.

Amino Acid Sequence↗

PubMed1996

Sequencing of a 17.6 kb segment on the right arm of yeast chromosome VII reveals 12 ORFs, including CCT, ADE3 and TR-I genes, homologues of the yeast PMT and EF1G genes, of the human and bacterial electron-transferring flavoproteins (beta-chain) and of the Escherichia coli phosphoserine phosphohydrolase, and five new ORFs.

A 17.6 kb DNA fragment from the right arm of chromosome VII of Saccharomyces cerevisiae has been sequenced and analysed. The sequence contains twelve open reading frames (ORFs) longer than 100 amino acids. Three genes had already been cloned and sequenced: CCT, ADE3 and TR-I. Two ORFs are similar to other yeast genes: G7722 with the YAL023 (PMT2) and PMT1 genes, encoding two integral membrane proteins, and G7727 with the first half of the genes encoding elongation factors 1gamma, TEF3 and TEF4. Two other ORFs, G7742 and G7744, are most probably yeast orthologues of the human and Paracoccus denitrificans electron-transferring flavoproteins (beta chain) and of the Escherichia coli phosphoserine phosphohydrolase. The five remaining identified ORFs do not show detectable homology with other protein sequences deposited in data banks. The sequence has been deposited in the EMBL data library under Accession Number Z49133.

Amino Acid Sequence↗

PubMed1994

Characteristics of biliary tract complications during laparoscopic cholecystectomy: a multi-institutional study.

We collected the records of 81 patients with biliary tract injuries occurring during laparoscopic cholecystectomy (LC) who were referred to 3 referral centers during a 33-month (May 1990 to March 1993) period. All records were reviewed to provide data concerning the anatomy of the lesion induced, method of injury, timing of injury detection, role of intraoperative cholangiography (IOC), methods of treatment, and outcome of these injuries. Injuries were classified by our own method as follows: (1) cystic duct leaks (n = 15), (2) bile leaks and/or ductal strictures (n = 27), and (3) ductal transections or excisions (n = 39). Peak occurrence by quarter of the year was 4th quarter, 1990 (Lahey), and 3rd quarter, 1991 (Mason), and 1st quarter, 1992 (Mayo). The majority (62%) of the injuries were recognized after LC. At the time of LC, 31 of 81 (38%) injuries were recognized and converted to open procedures. Data regarding IOC were available in 63 of 81 (78%) cases. In patients in whom IOC was not performed, 14 of 38 (37%) operations were converted; if an IOC was obtained and interpreted correctly, 13 of 21 (62%) operations were converted. Primary repair was attempted in 11 leaks and/or strictures, but 36% required additional treatment. Primary repair was used in six transections or excisions, and 17% have required further intervention. In patients who had biliary-enteric bypass (BEB) performed outside (17) versus at the referral institution (29), 94% (16 patients) versus 0%, respectively, required additional operative (e.g., revision of a hepaticojejunostomy) or nonoperative (e.g., radiologic or endoscopic stenting or balloon dilation) procedures. When used as initial therapy or after a primary ductal repair, stents (with or without balloon dilation) resolved 100% of simple cystic duct leaks and 91% of leaks and/or strictures. In conclusion, the peak incidence of LC-related biliary injuries appears to have passed. A completed and correctly interpreted IOC increases the chance of detection of biliary injuries intraoperatively and should assist surgeons who use routine IOC. Nonsurgical techniques allow treatment of most simple cystic duct leaks, major ductal leaks and/or strictures, and postoperative BEB strictures, although follow-up is limited. The poor results of pre-referral BEB is not surprising since all of these patients were selected for referral because their treatments had not been successful.

Bile Ducts↗

PubMed2001

Genetic and biochemical analyses of a eukaryotic-like phospholipase D of Pseudomonas aeruginosa suggest horizontal acquisition and a role for persistence in a chronic pulmonary infection model.

Phospholipases D (PLDs) are virtually ubiquitous in eukaryotic organisms; however, they are relatively uncommon in prokaryotes. In this report, we demonstrate that the environmentally acquired, opportunistic pathogen Pseudomonas aeruginosa expresses PLD activity. A gene designated pldA was identified in the genomic database of P. aeruginosa PAO1 encoding a protein with significant homology to eukaryotic PLDs, but not to any prokaryotic PLDs. PldA is most homologous to PLDs from mammals and yeast. The pldA gene was cloned and shown to express an approximately 116 kDa protein with calcium-regulated PLD activity that is localized to the periplasm. Interestingly, not all strains of P. aeruginosa carry pldA. When present, pldA is always linked to an open reading frame (ORF), ORF4, and a gene (vgrA1) encoding a protein homologous to Vgr from Escherichia coli. Vgr proteins contain regularly repeated dipeptide motifs (valine-glycine repeats). In E. coli, genes encoding Vgr are associated with multicopy genetic elements designated Rhs (rearrangement hot-spots). P. aeruginosa PAO1 has 10 vgr homologues dispersed throughout its genome, but the copy number of these genetic elements varies considerably in different strains. Neither vgrA1 nor ORF4 is present in strains lacking pldA. Furthermore, sequences flanking vgrA1, pldA and ORF4 in the P. aeruginosa strains examined are highly conserved, suggesting a specific site of insertion. These and other data suggest that vgrA1, pldA and ORF4 constitute an approximately 7 kb mobile genetic element and that pldA was acquired horizontally, perhaps from a eukaryotic organism. Competition studies between a PldA knock-out mutant and the parental wild-type strain indicate that PldA contributes to the ability of P. aeruginosa PAO1 to persist in a chronic pulmonary infection model in rats.

Animals↗

PubMed2004

The Aspergillus fumigatus transcriptional activator CpcA contributes significantly to the virulence of this fungal pathogen.

We have cloned and characterized the Aspergillus fumigatus cpcA gene encoding the transcriptional activator of the cross-pathway control system of amino acid biosynthesis. cpcA encodes a functional orthologue of Saccharomyces cerevisiae Gcn4p. The coding sequence of the 2.2 kb transcript is preceded by two short upstream open reading frames, the larger one being well conserved among Aspergilli. Deletion strains in which either the coding sequence or the entire locus are replaced by a bifunctional dominant marker are impaired in their cross-pathway control response upon amino acid starvation, as demonstrated by analyses of selected reporter genes and specific enzymatic activities. In a murine model of pulmonary aspergillosis, cpcAdelta strains display attenuated virulence. Pathogenicity is restored to wild-type levels in strains with reconstitution of the genomic locus. Competitive mixed infection experiments additionally demonstrate that cpcAdelta strains are less able to survive in vivo than their wild-type progenitor. Our data suggest that specific stress conditions are encountered by A. fumigatus within the mammalian host and that the fungal cross-pathway control system plays a significant role in pulmonary aspergillosis.

Amino Acid Sequence↗

PubMed1993

The nucleotide sequence of a 2.1 kb fragment from chromosome VI of Saccharomyces cerevisiae identifies a tRNA(Gly) gene, part of a delta element and a palindromic sequence.

The nucleotide sequence was determined of a 2.1 kb DNA fragment located at approximately 35 kb to the right of the centromere of chromosome VI from Saccharomyces cerevisiae. Analysis revealed the presence of a tRNA(GLy) gene, part of a delta element and a remarkable palindromic sequence. The longest open reading frame found encodes a putative protein of 195 amino acids. Although the fragment was isolated by hybridization to a human diacylglycerol kinase cDNA, no evidence was obtained for the presence of a gene encoding diacylglycerol kinase.

Base Sequence↗

PubMed1993

Isolation and characterization of a rice cDNA clone encoding ATP/ADP translocator.

We isolated a rice cDNA clone which encodes an open reading frame of 382 amino acids. Its deduced amino acid sequence corresponds to an ATP/ADP translocator protein. Its homology with a maize ATP/ADP translocator was 83.9% in nucleotide sequence, and 90.2% of the amino acid level. Expression of this gene is regulated by such external stresses as salinity and low temperature.

Amino Acid Sequence↗

PubMed1991

Cloning and nucleotide sequence of the chimpanzee c-myc gene.

A DNA fragment covering the chimpanzee c-myc locus was cloned from the DNA of peripheral blood lymphocytes, sequenced, and compared to its human c-myc counterpart. The two nucleotide sequences were found to be highly homologous (99%). The divergence rate between the two species was 0.4% in exons and 1.7% in introns. The different TATA-boxes described in the human myc gene were also identified in the chimpanzee sequence and an open reading frame (ORF) was observed which overlaps the chimpanzee c-myc first exon. This latter ORF contained three silent mutations with regard to the human region, whereas the chimpanzee Myc oncoprotein coded by exons 2 and 3 differed by two amino acids from the human one.

Animals↗