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At least 991 records · Page 55Linked to original sources
PubMed1993

The SCH9 protein kinase mRNA contains a long 5' leader with a small open reading frame.

The SCH9 yeast gene, that was previously identified as a suppressor of cdc25 and ras1- ras2-ts temperature-sensitive mutants, encodes a putative protein kinase that positively regulates the progression of yeast cells through the G1 phase of the cell cycle. We have determined the structure of the SCH9 transcription unit, using primer extension and S1 mapping techniques. The corresponding mRNA included an unusually long 5' region of more than 600 nucleotides preceding the major open reading frame (ORF). While the latter corresponded to a protein of 824 amino acids, an upstream open reading frame (uORF) within the 5' leader could potentially encode a 54 amino acid peptide. To investigate the role of the AUGs within the uORF, we engineered chimaeric plasmid vectors in which SCH9 sequences including the promoter, the mRNA leader and the first 514 nucleotides of the major ORF were fused in-frame with beta-galactosidase-coding sequences. Upon introduction into yeast cells, the fusion protein was efficiently expressed. However, mutational disruption of the uORF using oligonucleotide-directed mutagenesis did not affect the level of expression of the fusion protein. This indicates that regulatory mechanisms in Saccharomyces cerevisiae prevent upstream AUGs within the SCH9 mRNA leader sequence from influencing translation from downstream initiation codons.

Alleles↗
PubMed2001

Imprinting and evolution of two Kruppel-type zinc-finger genes, ZIM3 and ZNF264, located in the PEG3/USP29 imprinted domain.

We have isolated Kruppel-type (C2H2) zinc-finger genes, ZIM3 (zinc-finger gene 3 from imprinted domain) and ZNF264, located downstream of human and mouse USP29 genes (encoding ubiquitin-specific processing protease 29). In human, both ZIM3 and ZNF264 encode zinc-finger proteins with Kruppel-associated box (KRAB) A and B domains at the amino-terminal regions of the predicted proteins. In contrast, mouse Zim3 and Zfp264 seem to have lost protein-coding capability based on the lack of open reading frames (ORFs) in their cDNA sequences. In particular, the 3' end of the Zim3 transcript overlaps with the coding region of the adjacent gene Usp29 in an antisense orientation, indicating the conversion of mouse Zim3 into an antisense transcript gene for Usp29. The expression patterns of ZIM3 and ZNF264 have been largely conserved between human and mouse, with testis-specific expression of ZIM3 and ubiquitous expression of ZNF264, but high expression levels in adult testes in both species. Our studies also demonstrate that both mouse genes are imprinted with maternal expression of Zim3 in adult testes and paternal expression of Zfp264 in neonatal and adult brain. The reciprocal imprinting of two neighboring mouse genes, Zim3 and Zfp264, is consistent with a pattern observed frequently in other imprinted domains, and suggests that the imprinting of these two genes might be coregulated.

Amino Acid Sequence↗
PubMed2006

Evolution of Piperales--matK gene and trnK intron sequence data reveal lineage specific resolution contrast.

Piperales represent the largest basal angiosperm order with a nearly worldwide distribution. The order includes three species rich genera, Piper (ca. 2000 species), Peperomia (ca. 1500-1700 species), and Aristolochia s. l. (ca. 500 species). Sequences of the matK gene and the non-coding trnK group II intron are analysed for a dense set of 105 taxa representing all families (except Hydnoraceae) and all generic segregates (except Euglypha within Aristolochiaceae) of Piperales. A large number of highly informative indels are found in the Piperales trnK/matK dataset. Within a narrow region approximately 500 nt downstream in the matK coding region (CDS), a length variable simple sequence repeat (SSR) expansion segment occurs, in which insertions and deletions have led to short frame-shifts. These are corrected shortly afterwards, resulting in a maximum of six amino acids being affected. Furthermore, additional non-functional matK copies were found in Zippelia begoniifolia, which can easily be discriminated from the functional open reading frame (ORF). The trnK/matK sequence data fully resolve relationships within Peperomia, whereas they are not effective within Piper. The resolution contrast is correlated with the rate heterogeneity between those lineages. Parsimony, Bayesian and likelihood analyses result in virtually the same topology, and converge on the monophyly of Piperaceae and Saururaceae. Lactoris gains high support as sister to Aristolochiaceae subf. Aristolochioideae, but the different tree inference methods yield conflicting results with respect to the relationships of subfam. Asaroideae. In Piperaceae, a clade formed by the monotypic genus Zippelia and the small genus Manekia (=Sarcorhachis) is sister to the two large genera Piper and Peperomia.

Base Sequence↗
PubMed1999

Genetic analysis of the Serratia marcescens N28b O4 antigen gene cluster.

The Serratia marcescens N28b wbbL gene has been shown to complement the rfb-50 mutation of Escherichia coli K-12 derivatives, and a wbbL mutant has been shown to be impaired in O4-antigen biosynthesis (X. Rubirés, F. Saigí, N. Piqué, N. Climent, S. Merino, S. Albertí, J. M. Tomás, and M. Regué, J. Bacteriol. 179:7581-7586, 1997). We analyzed a recombinant cosmid containing the wbbL gene by subcloning and determination of O-antigen production phenotype in E. coli DH5alpha by sodium dodecyl sulfate-polyacrylamide electrophoresis and Western blot experiments with S. marcescens O4 antiserum. The results obtained showed that a recombinant plasmid (pSUB6) containing about 10 kb of DNA insert was enough to induce O4-antigen biosynthesis. The same results were obtained when an E. coli K-12 strain with a deletion of the wb cluster was used, suggesting that the O4 wb cluster is located in pSUB6. No O4 antigen was produced when plasmid pSUB6 was introduced in a wecA mutant E. coli strain, suggesting that O4-antigen production is wecA dependent. Nucleotide sequence determination of the whole insert in plasmid pSUB6 showed seven open reading frames (ORFs). On the basis of protein similarity analysis of the ORF-encoded proteins and analysis of the S. marcescens N28b wbbA insertion mutant and wzm-wzt deletion mutant, we suggest that the O4 wb cluster codes for two dTDP-rhamnose biosynthetic enzymes (RmlDC), a rhamnosyltransferase (WbbL), a two-component ATP-binding-cassette-type export system (Wzm Wzt), and a putative glycosyltransferase (WbbA). A sequence showing DNA homology to insertion element IS4 was found downstream from the last gene in the cluster (wbbA), suggesting that an IS4-like element could have been involved in the acquisition of the O4 wb cluster.

ATP-Binding Cassette Transporters↗
PubMed2003

Initiation of translation and cellular localization of Theileria annulata casein kinase IIalpha: implication for its role in host cell transformation.

Theileria annulata and T. parva are protozoa that infect bovine leukocytes which leads to subsequent transformation and uncontrolled proliferation of these cells. It has been proposed that the CKIIalpha subunit of T. parva induces mitogenic pathways of host leukocytes by being exported into the host cell. The evidence for this is the existence of a predicted N-terminal secretion signal-like peptide. We tested this hypothesis by analyzing gene structure, translation, and protein localization of the T. annulata CKIIalpha (TaCKIIalpha). The determined TaCKIIalpha-ORF potentially codes for a 50 kDa protein with an N-terminal extension including a possible signal sequence not present in CKIIalpha proteins of non-Theileria species. However, antisera raised against TaCKIIalpha recognized a protein of a molecular weight of about 40 kDa and, therefore, inconsistent with this predicted molecular weight. We demonstrate by in vitro transcription/translation that this discrepancy is due to translation from a downstream initiation site omitting the putative N-terminal signal sequence and thus excluding the notion that the protein product is secreted via the classical secretory pathway. In corroboration immunofluorescence investigations suggest that the TaCKIIalpha subunit is confined to the parasite schizonts within the host cell. On the basis of the above findings it seems highly unlikely that export via the classical pathway of the parasite CKIIalpha is the way in which this protein possibly contributes to host cell transformation.

Amino Acid Sequence↗
PubMed2004

A novel genetic locus outside the symbiotic island is required for effective symbiosis of Bradyrhizobium japonicum with soybean Glycine max.

In order to investigate the symbiotic interaction between soybean and Bradyrhizobium japonicum, TnphoA mutagenesis of the microsymbiont was performed. Mutant strain 2-10 was found to induce a strongly reduced number of ineffective nodules. Ultrastructural analysis of the soybean nodule central tissue revealed the presence of numerous starch granules and vacuoles in the infected cells. In addition, the number of symbiosomes was extremely low, indicating an impaired interaction between the plant and invading bacteria. Cloning and sequencing of the mutated DNA region uncovered four open reading frames (ORFs) lacking any data base similarities. ORFs srrA1 and srrA2, the 2-10 TnphoA insertion site, are encoded in the same reading frame. A 35-kDa expression product in Escherichia coli indicated the presence of a common protein, called SrrA (symbiotically relevant region) in B. japonicum 110spc4, encoded by combined srrA1 and srrA2 genes. The analysis of gene disruption mutants revealed that srrB and srrC were also required for effective symbiosis with soybeans. Further downstream the gene for a putative inner membrane protein (pipA) of unknown function was encoded on the opposite strand. Primer extension studies led to the conclusion that the organization of genes differed from the RhizoBase annotation in this particular region of B. japonicum USDA110.

Amino Acid Sequence↗
PubMed2006

A dicistronic gene pair within a cluster of "EF-hand" protein genes in the genomes of Drosophila species.

Androcam is a Drosophila melanogaster calmodulin-related protein that functions specifically in the testis. We show that the Acam gene is part of a cluster of three intronless genes arranged in a head-to-tail manner. The additional genes also encode calmodulin-related proteins with testis-specific transcription. Acam and the 5'-most gene (gene1) generate monocistronic transcripts. Surprisingly, the central gene (gene2) is transcribed only as a dicistronic transcript with Acam. A similar cluster is found in D. yakuba. In D. pseudoobscura, the cluster contains four genes: two Acam-type genes downstream of a single gene related to both gene1 and gene2 and a fourth weakly related gene. Nevertheless, the D. pseudoobscura cluster also generates a dicistronic transcript from a gene pair analogous to the gene2-Acam pair. A cotranscribed gene1/2-Acam gene pair may be the founding feature of this locus. Although Acam protein is present in D. melanogaster and D. pseudoobscura testes, cognate proteins for the gene1/2-type ORFs are not detectable by immunoblotting and mass spectrometry techniques.

Animals↗
PubMed1999

Characterization and sequencing of glutamine synthetase cDNA from liver of the ureotelic gulf toadfish (Opsanus beta).

The hepatic enzyme, glutamine synthetase (GSase) is a pivotal protein in the regulation of urea synthesis in fish. The sequence of the DNA encoding for GSase from liver of the ureotelic gulf toadfish (Opsanus beta) was analyzed through a suite of molecular techniques (including cDNA cloning, RACE PCR, and genomic PCR). An open reading frame (ORF) was identified in the cDNA sequence which codes for a protein of 394 amino acids with high identity (86%) to dogfish shark GSase. In the course of generating a suitable probe, a partial sequence was also obtained for horned shark GSase which also had high identity with the dogfish shark gene (93%). Like the dogfish shark GSase, the toadfish gene has two methionine translation initiation sites; the downstream site apparently codes for a cytoplasmic isozyme, while the upstream site adds an N-terminal peptide leader sequence of 23 amino acids to the 'cytoplasmic' protein. This leader sequence has characteristics consistent with a mitochondrial targeting peptide, including a cleavage recognition motif (Arg-X-Phe) and the apparent ability to form an amphiphathic helix. Northern analysis revealed that there is a single predominant transcript of approximately 2 kb in size. These results are consistent with the interpretation that in the gulf toadfish GSase cytoplasmic and mitochondrial isozymes are coded for by a single gene and mRNA transcript which is differentially translated at either initiation site. These results are discussed in the context of prior results for enzyme kinetic characteristics and urea synthesis/excretion physiology.

Amino Acid Sequence↗
PubMed1995

Intracellular localization of polypeptides encoded in mouse hepatitis virus open reading frame 1A.

We have investigated the intracellular localization of several of the proteolytic cleavage products derived from the 5' portion of mouse hepatitis virus (MHV) gene 1. Antisera UP1 recognizes the N-terminal ORF1a cleavage product p28. Immunofluorescent staining of cells with this antisera resulted in a diffuse punctate pattern of cytoplasmic staining, indicating that this protein is widely distributed in the cytoplasm. Immunofluorescent staining of infected cells with antisera which recognize polypeptides p240 and p290 stained discrete vesicular perinuclear structures suggesting that these proteins localized to the Golgi. This was confirmed by double immunofluorescent staining of BHK cells expressing the MHV receptor (BHK-R) with a Golgi specific antibody in addition to our anti-MHV ORF1a antibodies. Antisera UP102 recognizes p28 and the immediately downstream p65 gene product. Double immunofluorescent staining of MHV infected BHK-R cells with UP102 labeled discrete vesicular structures overlapping the Golgi complex. In addition there was punctate staining more widely distributed in the cytoplasm. The simplest explanation for this pattern is that p65 is also localized to the Golgi region of the cell, whereas p28 is more widespread. Plasmids containing the first 4.7 and 6.75 kb of ORF 1a have been expressed using the coupled vaccinia virus-T7 polymerase system. Images obtained by immunofluorescent staining of transfectants with our anti-ORF1a antisera are similar to those obtained during infection with A59. These studies indicate that the signals which direct p290 to the Golgi are likely contained between the C-terminus of p28 and ORF1a residue 1494.

Animals↗
PubMed1993

The single group-I intron in the chloroplast rrnL gene of Chlamydomonas humicola encodes a site-specific DNA endonuclease (I-ChuI).

The single group-I intron (ChLSU.1) in the chloroplast (cp) large subunit rRNA-encoding gene (rrnL) of the green alga Chlamydomonas humicola is located at a position at which no introns have previously been characterized in other systems. In the present study, the nucleotide (nt) sequence of this 1118-bp intron was found to contain an internal open reading frame (ORF) that potentially encodes a basic protein of 218 amino acid residues. The putative C. humicola protein features two copies of the LAGLI-DADG motif and is part of the family of intron-encoded proteins comprising the endonucleases (ENases), I-SceI, I-SceIV and I-CsmI. Expression of the ChLSU.1 intron ORF in vitro in the presence of a 260-bp DNA fragment containing the exon 1-2 junction of an intronless version of the C. humicola rrnL resulted in specific cleavage of the DNA fragment very close to the intron insertion site. This novel intron-encoded ENase, designated I-ChuI, was also shown to generate a staggered cut with 4-nt (CTCG) 3'-OH overhangs 2 bp downstream from the intron insertion site.

Amino Acid Sequence↗
PubMed1994

Organization of the hydrogenase gene cluster from Bradyrhizobium japonicum: sequences and analysis of five more hydrogenase-related genes.

Previously, the deletion of a 2.9-kb chromosomal EcoRI fragment of DNA located 2.2 kb downstream from the end of the Bradyrhizobium japonicum hydrogenase structural genes caused lack of normal-sized hydrogenase (Hup) subunits and complete loss of Hup activity. It was suggested that this region encodes one or more genes required for Hup processing. Sequencing of a 3322-bp XcmI fragment of DNA covering this 2.9-kb EcoRI fragment within the hup gene cluster revealed the presence of five open reading frames (ORFs) designated hupG, hupH, hupI, hupJ and hupK, encoding polypeptides with calculated molecular masses of 15.8, 30.7, 7.6, 18.1 and 38 kDa, respectively. Based on deduced amino acid (aa) sequences, all five products of the hupGHIJK genes showed significant homology with other genes' products in several H2-utilizing bacteria. Of particular interest are HupG and HupI. HupG showed 70% similarity (28% identity) to the HyaE of the Escherichia coli hydrogenase-1 operon which was demonstrated to be involved in the processing of hydrogenase-1. HupI showed strong identity to rubredoxin and rubredoxin-like proteins from many other bacteria. The latter proteins contain two 'C-X-X-C' motifs, which may serve as iron ligands for non-heme iron proteins involved as intermediate electron carriers or in the assembly process for Fe-S (or NiFe-S) clusters.

Amino Acid Sequence↗
PubMed2004

SCC-112, a novel cell cycle-regulated molecule, exhibits reduced expression in human renal carcinomas.

We report here the identification and an initial characterization of a novel cell cycle-regulated molecule, SCC-112. SCC-112 cDNA (6744 bp) encodes a longest open reading frame (ORF) comprised of 1297 amino acids, representing a approximately 150-kDa nuclear protein. SCC-112 mRNA and protein levels were relatively high during the G2/M phase of the cell cycle in MDA-MB 435 breast cancer cells. Transient expression of SCC-112 cDNA in COS-1 cells led to an increase in the number of cells in sub-G1 phase and enhanced activity of caspase-3, a downstream effector of apoptosis. Stable transfection of SCC-112 cDNA in MDA-MB 231 breast cancer cells also led to an increase in the number of cells in sub-G1 phase ( approximately 2-3-fold), indicative of apoptosis. The examination of the paired sets of human normal and tumor tissues revealed that the SCC-112 mRNA level was significantly high in normal breast and kidney tissues as compared to the corresponding primary tumor tissues (P<0.0001; breast, n=50, and kidney, n=20). Consistent with these observations, SCC-112 protein expression (150 kDa) was high in a majority of the normal renal tissues examined as compared to the matched renal tumor tissues (67%, 1.2-fold to>10-fold, n=18). Taken together, these findings suggest that the SCC-112 gene expression is likely to be associated with normal cell growth and proliferation.

Adult↗
PubMed2002

The superantigen gene ypm is located in an unstable chromosomal locus of Yersinia pseudotuberculosis.

Yersinia pseudotuberculosis produces YPM (Y. pseudotuberculosis-derived mitogen), a superantigenic toxin that exacerbates the virulence of the bacterium in vivo. To date, three alleles of the superantigen gene (ypmA, ypmB, and ypmC) have been described. These genes are not found in all Y. pseudotuberculosis strains and have a low GC content, suggesting their location on mobile genetic elements. To elucidate this question, the genetic environment of the superantigen-encoding genes was characterized and 11 open reading frames (ORFs) were defined. Sequence analysis revealed that the ypm genes were not associated with plasmids, phages, transposons, or pathogenicity islands and that the superantigen genes were always located in the chromosome between ORF3 and ORF4. Nonsuperantigenic strains exhibited the same genetic organization of the locus but lacked the ypm gene between ORF3 and ORF4. A new insertion sequence, designated IS1398, which displays features of the Tn3 family, was characterized downstream of the ypmA and ypmC genes. A 13.3-kb region containing the ypm genes was not found in the genome of Y. pestis (CO92 and KIM 5 strains). We experimentally induced deletion of the ypm gene from a superantigen-expressing Y. pseudotuberculosis: using the association of aph(3')-IIIa and sacB genes, we demonstrated that when these reporter genes were present in the ypm locus, deletion of these genes was about 250 times more frequent than when they were located in another region of the Y. pseudotuberculosis chromosome. These results indicate that unlike other superantigenic toxin genes, the Yersinia ypm genes are not associated with mobile genetic elements but are inserted in an unstable locus of the genome.

Alleles↗
PubMed1995

An archaeal gene upstream of grpE different from eubacterial counterparts.

In some eubacteria with a dnaK locus in which grpE is close upstream of dnaK, grpE is preceded by an open reading frame (orf) believed to be a heat-shock gene. We also found an orf, orf16, upstream of grpE in the archaeon Methanosarcina mazei S-6, but this gene differs from the eubacterial counterpart: it is shorter, does not respond to a temperature upshift as heat-shock genes do, and the deduced protein Orf16, does not resemble the proteins coded by the eubacterial equivalents. orf16 is expressed monocistronically, with a transcription initiation site 24 bases upstream of the translation start codon, 22 bases downstream of a putative promoter identical to the consensus promoter for genes in methanogens. This initiation site is used by heat-shocked and non-heat-shocked cells in the two morphologic stages of M. mazei S-6 tested, i.e., packets and single cells. Three transcription termination sites were identified, one of which is detectable only in non-heat-shocked cells. Data from comparative analyses of the Orf16 deduced amino acid sequence and those of other known proteins, as well as the apparent biochemical characteristics of Orf16, suggest that the latter is a membrane molecule.

Amino Acid Sequence↗
PubMed1996

Nucleotide sequence and role in DNA amplification of the direct repeats composing the amplifiable element AUD1 of Streptomyces lividans 66.

The amplifiable unit of DNA no. 1 (AUD1) of Streptomyces lividans consists of three 1 kb repeats (left direct repeat, LDR; middle direct repeat, MDR; and the slightly different right direct repeat, RDR) and two 4.7 kb repeats alternately arranged in identical orientation to each other. Both 4.7 kb repeats have been sequenced. They are identical and contain one open reading frame (orf4.7). The deduced amino acid sequence has a low similarity to chitinases, and two amino acid repeats present high similarities to fibronectin type III modules. Sequencing had previously shown that the ORF corresponding to each 1 kb repeat encodes a putative DNA-binding protein. Crude extracts of Escherichia coli overexpressing the orfRDR-encoded protein and of S. lividans Jni1, having a high amplification of AUD1 and therefore orfMDR, were used in gel retardation assays. The orfRDR- and probably the orfMDR-encoded proteins can bind to an imperfect palindromic sequence upstream from MDR and RDR and to another sequence downstream from RDR. An extrachromosomal DNA amplification system was constructed containing different combinations of the sequences composing AUD1. In mutants having a deletion of the chromosomal AUD1, the 4.7 kb repeats could be reduced in size, mutated or replaced by E. coli DNA without altering the ability to amplify when RDR was present. Therefore, the only function of the 4.7 kb repeats in amplification is to provide directly repeated DNA sequences. When RDR was lacking or mutated, no amplification was observed. This strongly suggests that the DNA-binding protein encoded by orfRDR is required for AUD1 amplification.

Amino Acid Sequence↗
PubMed2005

Molecular characterization of a multidrug-resistant strain of enteroinvasive Escherichia coli O164 isolated in Japan.

Enteroinvasive Escherichia coli (EIEC) O164 strain RIMD05091045 was isolated from a travelling patient suffering from diarrhoea at the Osaka airport quarantine facility in Japan. The strain showed multidrug resistance against streptomycin, spectinomycin, co-trimoxazole (trimethoprim/sulfamethoxazole) and ampicillin, and reduced susceptibility to ciprofloxacin. Molecular characterization of the multidrug-resistance phenotype revealed the presence of a class 1 integron containing three genes, a dihydrofolate reductase type XII gene, dfrXII, which confers resistance to trimethoprim, an aminoglycoside adenyltransferase gene, aadA2, which confers resistance to streptomycin and spectinomycin, and an ORF of unknown function. Southern blot hybridization and conjugation experiments showed that the class 1 integron was located on a transferable plasmid that was less than 90 kb in size. The resistance of EIEC O164 to ampicillin was found to be due to the presence of TEM-1 beta-lactamase. On the other hand, a single mutation that has not previously been described, P158-to-S, was detected downstream of the quinolone-resistance-determining region of parC of topoisomerase IV and may be responsible for the reduced susceptibility to ciprofloxacin in this strain.

Ampicillin Resistance↗
PubMed2002

Degradation of aromatics and chloroaromatics by Pseudomonas sp. strain B13: cloning, characterization, and analysis of sequences encoding 3-oxoadipate:succinyl-coenzyme A (CoA) transferase and 3-oxoadipyl-CoA thiolase.

3-oxoadipate:succinyl-coenzyme A (CoA) transferase and 3-oxoadipyl-CoA thiolase carry out the ultimate steps in the conversion of benzoate and 3-chlorobenzoate to tricarboxylic acid cycle intermediates in bacteria utilizing the 3-oxoadipate pathway. This report describes the characterization of DNA fragments with the overall length of 5.9 kb from Pseudomonas sp. strain B13 that encode these enzymes. DNA sequence analysis revealed five open reading frames (ORFs) plus an incomplete one. ORF1, of unknown function, has a length of 414 bp. ORF2 (catI) encodes a polypeptide of 282 amino acids and starts at nucleotide 813. ORF3 (catJ) encodes a polypeptide of 260 amino acids and begins at nucleotide 1661. CatI and CatJ are the subunits of the 3-oxoadipate:succinyl-CoA transferase, whose activity was demonstrated when both genes were ligated into expression vector pET11a. ORF4, termed catF, codes for a protein of 401 amino acid residues with a predicted mass of 41,678 Da with 3-oxoadipyl-CoA thiolase activity. The last three ORFs seem to form an operon since they are oriented in the same direction and showed an overlapping of 1 bp between catI and catJ and of 4 bp between catJ and catF. Conserved functional groups important for the catalytic activity of CoA transferases and thiolases were identified in CatI, CatJ, and CatF. ORF5 (catD) encodes the 3-oxoadipate enol-lactone hydrolase. An incomplete ORF6 of 1,183 bp downstream of ORF5 and oriented in the opposite direction was found. The protein sequence deduced from ORF6 showed a putative AMP-binding domain signature.

Acetyl-CoA C-Acyltransferase↗
PubMed1992

Convergent and overlapping transcripts of the Chlamydia trachomatis 7.5-kb plasmid.

Transcription of the 7.5-kb cryptic plasmid of Chlamydia trachomatis serovar L2 was investigated. Faint, diffuse transcripts of about 1.6 and 2.2 kb and intense short transcripts of about 250 and 430 bases were identified by Northern blot analysis. The short transcripts were found to have a common 5' end corresponding to bp 501 relative to the unique BamHI site of the plasmid and to terminate at different downstream sites. Putative promoter sequences of TTGCCA and TATATT, which closely resemble the consensus recognition site of Escherichia coli sigma 70, were identified at the -35 and -10 positions upstream from the 5' end of the short transcripts made in chlamydia. Transcripts of similar sizes were also expressed from this promoter in E. coli harboring a recombinant plasmid encoding the short transcripts. The short transcripts encode a common open reading frame (ORF) of 34 codons; however, a strong ribosome binding site was not found in the vicinity of the initiator codon, and it is not known whether the transcripts are translated in vivo. A large ORF of 330 codons, which has been shown to encode a hypothetical protein containing conserved domains of recombinase-like proteins, is antisense to the short transcripts. Transcripts encoding the large ORF could not be detected directly by Northern blot or primer extension analysis. However, transcripts were detected by polymerase chain reaction amplification of the large ORF cDNA and when Southern blots of single-stranded antisense DNA for the large ORF were probed with radiolabeled RNA synthesized by host-free chlamydial reticulate bodies. Thus, both strands of the chlamydial plasmid are transcribed in the region encoding the short transcripts. We propose that the short transcripts play a regulatory role as antisense RNAs.

Amino Acid Sequence↗