scholarly journals Transcription and Regulation of the Bidirectional Hydrogenase in the Cyanobacterium Nostoc sp. Strain PCC 7120

2007 ◽  
Vol 73 (17) ◽  
pp. 5435-5446 ◽  
Author(s):  
Johannes Sjöholm ◽  
Paulo Oliveira ◽  
Peter Lindblad
Keyword(s):  
Open Reading Frames ◽  
Start Codon ◽  
Hydrogenase Activity ◽  
Promoter Regions ◽  
Mobility Shift ◽  
Reading Frame ◽  
Genes Encoding ◽  
Lexa Binding ◽  
Bidirectional Hydrogenase ◽  
Pcc 7120

ABSTRACT The filamentous, heterocystous cyanobacterium Nostoc sp. strain PCC 7120 (Anabaena sp. strain PCC 7120) possesses an uptake hydrogenase and a bidirectional enzyme, the latter being capable of catalyzing both H2 production and evolution. The completely sequenced genome of Nostoc sp. strain PCC 7120 reveals that the five structural genes encoding the bidirectional hydrogenase (hoxEFUYH) are separated in two clusters at a distance of approximately 8.8 kb. The transcription of the hox genes was examined under nitrogen-fixing conditions, and the results demonstrate that the cluster containing hoxE and hoxF can be transcribed as one polycistronic unit together with the open reading frame alr0750. The second cluster, containing hoxU, hoxY, and hoxH, is transcribed together with alr0763 and alr0765, located between the hox genes. Moreover, alr0760 and alr0761 form an additional larger operon. Nevertheless, Northern blot hybridizations revealed a rather complex transcription pattern in which the different hox genes are expressed differently. Transcriptional start points (TSPs) were identified 66 and 57 bp upstream from the start codon of alr0750 and hoxU, respectively. The transcriptions of the two clusters containing the hox genes are both induced under anaerobic conditions concomitantly with the induction of a higher level of hydrogenase activity. An additional TSP, within the annotated alr0760, 244 bp downstream from the suggested translation start codon, was identified. Electrophoretic mobility shift assays with purified LexA from Nostoc sp. strain PCC 7120 demonstrated specific interactions between the transcriptional regulator and both hox promoter regions. However, when LexA from Synechocystis sp. strain PCC 6803 was used, the purified protein interacted only with the promoter region of the alr0750-hoxE-hoxF operon. A search of the whole Nostoc sp. strain PCC 7120 genome demonstrated the presence of 216 putative LexA binding sites in total, including recA and recF. This indicates that, in addition to the bidirectional hydrogenase gene, a number of other genes, including open reading frames connected to DNA replication, recombination, and repair, may be part of the LexA regulatory network in Nostoc sp. strain PCC 7120.

scholarly journals RegA, an AraC-Like Protein, Is a Global Transcriptional Regulator That Controls Virulence Gene Expression in Citrobacter rodentium

2008 ◽  
Vol 76 (11) ◽  
pp. 5247-5256 ◽  
Author(s):  
Emily Hart ◽  
Ji Yang ◽  
Marija Tauschek ◽  
Michelle Kelly ◽  
Matthew J. Wakefield ◽  
...  
Keyword(s):  
Mammalian Cells ◽  
Virulence Gene ◽  
Virulence Determinants ◽  
Intestinal Colonization ◽  
Citrobacter Rodentium ◽  
Promoter Regions ◽  
Reading Frame ◽  
Virulence Gene Expression ◽  
Genes Encoding ◽  
Colonization Factors

ABSTRACT Citrobacter rodentium is an attaching and effacing pathogen which causes transmissible colonic hyperplasia in mice. Infection with C. rodentium serves as a model for infection of humans with enteropathogenic and enterohemorrhagic Escherichia coli. To identify novel colonization factors of C. rodentium, we screened a signature-tagged mutant library of C. rodentium in mice. One noncolonizing mutant had a single transposon insertion in an open reading frame (ORF) which we designated regA because of its homology to genes encoding members of the AraC family of transcriptional regulators. Deletion of regA in C. rodentium resulted in markedly reduced colonization of the mouse intestine. Examination of lacZ transcriptional fusions using promoter regions of known and putative virulence-associated genes of C. rodentium revealed that RegA strongly stimulated transcription of two newly identified genes located close to regA, which we designated adcA and kfcC. The cloned adcA gene conferred autoaggregation and adherence to mammalian cells to E. coli strain DH5α, and a kfc mutation led to a reduction in the duration of intestinal colonization, but the kfc mutant was far less attenuated than the regA mutant. These results indicated that other genes of C. rodentium whose expression required activation by RegA were required for colonization. Microarray analysis revealed a number of RegA-regulated ORFs encoding proteins homologous to known colonization factors. Transcription of these putative virulence determinants was activated by RegA only in the presence of sodium bicarbonate. Taken together, these results show that RegA is a global regulator of virulence in C. rodentium which activates factors that are required for intestinal colonization.

scholarly journals FuncPEP: A Database of Functional Peptides Encoded by Non-Coding RNAs

2020 ◽  
Vol 6 (4) ◽  
pp. 41
Author(s):  
Mihnea P. Dragomir ◽  
Ganiraju C. Manyam ◽  
Leonie Florence Ott ◽  
Léa Berland ◽  
Erik Knutsen ◽  
...  
Keyword(s):  
Open Reading Frames ◽  
Start Codon ◽  
Small Peptides ◽  
Reading Frame ◽  
Cellular Processes ◽  
New Class ◽  
Web Resource ◽  
Non Coding Rnas ◽  
Functional Peptides ◽  
Small Open Reading Frames

Non-coding RNAs (ncRNAs) are essential players in many cellular processes, from normal development to oncogenic transformation. Initially, ncRNAs were defined as transcripts that lacked an open reading frame (ORF). However, multiple lines of evidence suggest that certain ncRNAs encode small peptides of less than 100 amino acids. The sequences encoding these peptides are known as small open reading frames (smORFs), many initiating with the traditional AUG start codon but terminating with atypical stop codons, suggesting a different biogenesis. The ncRNA-encoded peptides (ncPEPs) are gradually becoming appreciated as a new class of functional molecules that contribute to diverse cellular processes, and are deregulated in different diseases contributing to pathogenesis. As multiple publications have identified unique ncPEPs, we appreciated the need for assembling a new web resource that could gather information about these functional ncPEPs. We developed FuncPEP, a new database of functional ncRNA encoded peptides, containing all experimentally validated and functionally characterized ncPEPs. Currently, FuncPEP includes a comprehensive annotation of 112 functional ncPEPs and specific details regarding the ncRNA transcripts that encode these peptides. We believe that FuncPEP will serve as a platform for further deciphering the biologic significance and medical use of ncPEPs. The link for FuncPEP database can be found at the end of the Introduction Section.

scholarly journals Identification of Chlamydia trachomatis Genomic Sequences Recognized by Chlamydial Divalent Cation-Dependent Regulator A (DcrA)

2005 ◽  
Vol 187 (2) ◽  
pp. 443-448 ◽  
Author(s):  
Annette Rau ◽  
Susan Wyllie ◽  
Judy Whittimore ◽  
Jane E. Raulston
Keyword(s):  
Chlamydia Trachomatis ◽  
Divalent Cation ◽  
Open Reading Frames ◽  
Genomic Sequences ◽  
Mobility Shift ◽  
Reading Frame ◽  
E Coli ◽  
Resultant Data ◽  
Mobility Shift Assay ◽  
Titration Assay

ABSTRACT The Chlamydia trachomatis divalent cation-dependent regulator (DcrA), encoded by open reading frame CT296, is a distant relative of the ferric uptake regulator (Fur) family of iron-responsive regulators. Chlamydial DcrA specifically binds to a consensus Escherichia coli Fur box and is able to complement an E. coli Fur mutant. In this report, the E. coli Fur titration assay (FURTA) was used to locate chlamydial genomic sequences that are recognized by E. coli Fur. The predictive regulatory regions of 28 C. trachomatis open reading frames contained sequences functionally recognized by E. coli Fur; targets include components of the type III secretion pathway, elements involved in envelope and cell wall biogenesis, predicted transport proteins, oxidative defense enzymes, and components of metabolic pathways. Selected FURTA-positive sequences were subsequently examined for recognition by C. trachomatis DcrA using an electrophoretic mobility shift assay. The resultant data show that C. trachomatis DcrA binds to native chlamydial genomic sequences and, overall, substantiate a functional relationship between chlamydial DcrA and the Fur family of regulators.

scholarly journals Genetic Composition of the Bacillus subtilis SOS System

2005 ◽  
Vol 187 (22) ◽  
pp. 7655-7666 ◽  
Author(s):  
Nora Au ◽  
Elke Kuester-Schoeck ◽  
Veena Mandava ◽  
Laura E. Bothwell ◽  
Susan P. Canny ◽  
...  
Keyword(s):  
Dna Damage ◽  
Bacillus Subtilis ◽  
Binding Sites ◽  
Consensus Sequence ◽  
Transcriptional Response ◽  
Open Reading Frames ◽  
Mobility Shift ◽  
E Coli ◽  
Recombination Protein ◽  
Lexa Binding

ABSTRACT The SOS response in bacteria includes a global transcriptional response to DNA damage. DNA damage is sensed by the highly conserved recombination protein RecA, which facilitates inactivation of the transcriptional repressor LexA. Inactivation of LexA causes induction (derepression) of genes of the LexA regulon, many of which are involved in DNA repair and survival after DNA damage. To identify potential RecA-LexA-regulated genes in Bacillus subtilis, we searched the genome for putative LexA binding sites within 300 bp upstream of the start codons of all annotated open reading frames. We found 62 genes that could be regulated by putative LexA binding sites. Using mobility shift assays, we found that LexA binds specifically to DNA in the regulatory regions of 54 of these genes, which are organized in 34 putative operons. Using DNA microarray analyses, we found that 33 of the genes with LexA binding sites exhibit RecA-dependent induction by both mitomycin C and UV radiation. Among these 33 SOS genes, there are 22 distinct LexA binding sites preceding 18 putative operons. Alignment of the distinct LexA binding sites reveals an expanded consensus sequence for the B. subtilis operator: 5′-CGAACATATGTTCG-3′. Although the number of genes controlled by RecA and LexA in B. subtilis is similar to that of Escherichia coli, only eight B. subtilis RecA-dependent SOS genes have homologous counterparts in E. coli.

scholarly journals Transcriptional and Functional Analysis of the Neisseria gonorrhoeae Fur Regulon

2009 ◽  
Vol 192 (1) ◽  
pp. 77-85 ◽  
Author(s):  
Lydgia A. Jackson ◽  
Thomas F. Ducey ◽  
Michael W. Day ◽  
Jeremy B. Zaitshik ◽  
Joshua Orvis ◽  
...  
Keyword(s):  
Neisseria Gonorrhoeae ◽  
Transcriptional Control ◽  
Cell Communication ◽  
Essential Element ◽  
Open Reading Frames ◽  
Promoter Regions ◽  
Mobility Shift ◽  
Sequence Logos ◽  
Mobility Shift Assay

ABSTRACT To ensure survival in the host, bacteria have evolved strategies to acquire the essential element iron. In Neisseria gonorrhoeae, the ferric uptake regulator Fur regulates metabolism through transcriptional control of iron-responsive genes by binding conserved Fur box (FB) sequences in promoters during iron-replete growth. Our previous studies showed that Fur also controls the transcription of secondary regulators that may, in turn, control pathways important to pathogenesis, indicating an indirect role for Fur in controlling these downstream genes. To better define the iron-regulated cascade of transcriptional control, we combined three global strategies—temporal transcriptome analysis, genomewide in silico FB prediction, and Fur titration assays (FURTA)—to detect genomic regions able to bind Fur in vivo. The majority of the 300 iron-repressed genes were predicted to be of unknown function, followed by genes involved in iron metabolism, cell communication, and intermediary metabolism. The 107 iron-induced genes encoded hypothetical proteins or energy metabolism functions. We found 28 predicted FBs in FURTA-positive clones in the promoters and within the open reading frames of iron-repressed genes. We found lower levels of conservation at critical thymidine residues involved in Fur binding in the FB sequence logos of FURTA-positive clones with intragenic FBs than in the sequence logos generated from FURTA-positive promoter regions. In electrophoretic mobility shift assay studies, intragenic FBs bound Fur with a lower affinity than intergenic FBs. Our findings further indicate that transcription under iron stress is indirectly controlled by Fur through 12 potential secondary regulators.

scholarly journals Identification of an ATP-Driven, Osmoregulated Glycine Betaine Transport System in Listeria monocytogenes

1999 ◽  
Vol 65 (9) ◽  
pp. 4040-4048 ◽  
Author(s):  
Rinkei Ko ◽  
Linda Tombras Smith
Keyword(s):  
Listeria Monocytogenes ◽  
Glycine Betaine ◽  
Open Reading Frames ◽  
Transport Systems ◽  
High Concentration ◽  
Promoter Regions ◽  
Reading Frame ◽  
Transposon Insertion ◽  
Food Borne ◽  
Reading Frames

ABSTRACT The ability of the gram-positive, food-borne pathogenListeria monocytogenes to tolerate environments of elevated osmolarity and reduced temperature is due in part to the transport and accumulation of the osmolyte glycine betaine. Previously we showed that glycine betaine transport was the result of Na+-glycine betaine symport. In this report, we identify a second glycine betaine transporter from L. monocytogenes which is osmotically activated but does not require a high concentration of Na+ for activity. By using a pool of Tn917-LTV3 mutants, a salt- and chill-sensitive mutant which was also found to be impaired in its ability to transport glycine betaine was isolated. DNA sequence analysis of the region flanking the site of transposon insertion revealed three open reading frames homologous to opuA from Bacillus subtilis and proU from Escherichia coli, both of which encode glycine betaine transport systems that belong to the superfamily of ATP-dependent transporters. The three open reading frames are closely spaced, suggesting that they are arranged in an operon. Moreover, a region upstream from the first reading frame was found to be homologous to the promoter regions of both opuAand proU. One unusual feature not shared with these other two systems is that the start codons for two of the open reading frames in L. monocytogenes appear to be TTG. That glycine betaine uptake is nearly eliminated in the mutant strain when it is assayed in the absence of Na+ is an indication that only the ATP-dependent transporter and the Na+-glycine betaine symporter occur in L. monocytogenes.

scholarly journals Organization and Expression of a Thermus thermophilusArginine Cluster: Presence of Unidentified Open Reading Frames and Absence of a Shine-Dalgarno Sequence

2000 ◽  
Vol 182 (20) ◽  
pp. 5911-5915 ◽  
Author(s):  
Rony Sanchez ◽  
Martine Roovers ◽  
Nicolas Glansdorff
Keyword(s):  
Specific Binding ◽  
Thermus Thermophilus ◽  
Enteric Bacteria ◽  
Open Reading Frames ◽  
Start Codon ◽  
Mobility Shift ◽  
Dnase I Footprinting ◽  
Shine Dalgarno Sequence ◽  
Related Metabolite ◽  
Reading Frames

ABSTRACT A group of genes regulated by arginine was found clustered in the order argF-ORF1-argC-argJ-ORF4 between other, as yet uncharacterized, open reading frames (ORFs). Transcription starts were identified immediately upstream fromargF and ORF4. Arginine repressed transcription that was initiated at argF but induced transcription of ORF4. The functions of ORF1 and ORF4 are unknown, but analysis of the sequence of ORF4 suggests that it is a membrane protein, possibly involved in transport of arginine or a related metabolite. Mobility shift and DNase I footprinting have revealed specific binding of pure Escherichia coli ArgR to the promoter region of Thermus thermophilus argF. These results suggest that argF transcription is controlled by a repressor homologous to those characterized in enteric bacteria and bacilli. Thermus argF mRNA is devoid of Shine-Dalgarno (SD) sequences. However, downstream from the ATG start codon ofargF and many other Thermus genes (with or without an SD box), sequences were found to be complementary to nucleotides 1392 to 1409 of Thermus 16S rRNA, suggesting that an mRNA-rRNA base pairing in this region is important for correct translation initiation.

scholarly journals Characterization of the CpxRA Regulon in Haemophilus ducreyi

2010 ◽  
Vol 78 (11) ◽  
pp. 4779-4791 ◽  
Author(s):  
Maria Labandeira-Rey ◽  
Chad A. Brautigam ◽  
Eric J. Hansen
Keyword(s):  
Stress Response ◽  
Response Regulator ◽  
Cell Envelope ◽  
Bacterial Species ◽  
Regulatory System ◽  
Promoter Regions ◽  
Mobility Shift ◽  
Envelope Stress ◽  
Two Component ◽  
Genes Encoding

ABSTRACT The H aemophilus ducreyi 35000HP genome encodes a homolog of the CpxRA two-component cell envelope stress response system originally characterized in E scherichia coli. CpxR, the cytoplasmic response regulator, was shown previously to be involved in repression of the expression of the lspB-lspA2 operon (M. Labandeira-Rey, J. R. Mock, and E. J. Hansen, Infect. Immun. 77:3402-3411, 2009). In the present study, the H. ducreyi CpxR and CpxA proteins were shown to closely resemble those of other well-studied bacterial species. A cpxA deletion mutant and a CpxR-overexpressing strain were used to explore the extent of the CpxRA regulon. DNA microarray and real-time reverse transcriptase (RT) PCR analyses indicated several potential regulatory targets for the H. ducreyi CpxRA two-component regulatory system. Electrophoretic mobility shift assays (EMSAs) were used to prove that H. ducreyi CpxR interacted with the promoter regions of genes encoding both known and putative virulence factors of H. ducreyi, including the lspB-lspA2 operon, the flp operon, and dsrA. Interestingly, the use of EMSAs also indicated that H. ducreyi CpxR did not bind to the promoter regions of several genes predicted to encode factors involved in the cell envelope stress response. Taken together, these data suggest that the CpxRA system in H. ducreyi, in contrast to that in E. coli, may be involved primarily in controlling expression of genes not involved in the cell envelope stress response.

scholarly journals Unusual Organization for Lactose and Galactose Gene Clusters in Lactobacillus helveticus

2003 ◽  
Vol 69 (6) ◽  
pp. 3238-3243 ◽  
Author(s):  
Maria Grazia Fortina ◽  
Giovanni Ricci ◽  
Diego Mora ◽  
Simone Guglielmetti ◽  
Pier Luigi Manachini
Keyword(s):  
Regulatory Gene ◽  
Gene Clusters ◽  
Lactobacillus Helveticus ◽  
Open Reading Frames ◽  
Open Reading Frame ◽  
Transcriptional Level ◽  
Reading Frame ◽  
Genes Encoding ◽  
Lactose Utilization ◽  
Inducible Genes

ABSTRACT The nucleotide sequences of the Lactobacillus helveticus lactose utilization genes were determined, and these genes were located and oriented relative to one another. The lacLM genes (encoding the β-galactosidase protein) were in a divergent orientation compared to lacR (regulatory gene) and lacS (lactose transporter). Downstream from lacM was an open reading frame (galE) encoding a UDP-galactose 4 epimerase, and the open reading frame had the same orientation as lacM. The lacR gene was separated from the downstream lacS gene by 2.0 kb of DNA containing several open reading frames that were derived from fragmentation of another permease gene (lacS′). Northern blot analysis revealed that lacL, lacM, and galE made up an operon that was transcribed in the presence of lactose from an upstream lacL promoter. The inducible genes lacL and lacM were regulated at the transcriptional level by the LacR repressor. In the presence of glucose and galactose galE was transcribed from its promoter, suggesting that the corresponding enzyme can be expressed constitutively. Lactose transport was inducible by addition of lactose to the growth medium.

scholarly journals A Coding Sequence-Embedded Principle Governs Translational Reading Frame Fidelity

Research ◽  
2018 ◽  
Vol 2018 ◽  
pp. 1-15 ◽  
Author(s):  
Ji Wan ◽  
Xiangwei Gao ◽  
Yuanhui Mao ◽  
Xingqian Zhang ◽  
Shu-Bing Qian
Keyword(s):  
18S Rrna ◽  
Ribosome Profiling ◽  
Open Reading Frames ◽  
Start Codon ◽  
Data Sets ◽  
Protein Coding ◽  
Reading Frame ◽  
Codon Composition ◽  
Sequence Elements ◽  
Correct Reading

Upon initiation at a start codon, the ribosome must maintain the correct reading frame for hundreds of codons in order to produce functional proteins. While some sequence elements are able to trigger programmed ribosomal frameshifting (PRF), very little is known about how the ribosome normally prevents spontaneous frameshift errors that can have dire consequences if uncorrected. Using high resolution ribosome profiling data sets, we discovered that the translating ribosome uses the 3′ end of 18S rRNA to scan the AUG-like codons after the decoding process. The postdecoding mRNA:rRNA interaction not only contributes to predominant translational pausing, but also provides a retrospective mechanism to safeguard the ribosome in the correct reading frame. Partially eliminating the AUG-like “sticky” codons in the reporter message leads to increased +1 frameshift errors. Remarkably, mutating the highly conserved CAU triplet of 18S rRNA globally changes the codon “stickiness”. Further supporting the role of “sticky” sequences in reading frame maintenance, the codon composition of open reading frames is highly optimized across eukaryotic genomes. These results suggest an important layer of information embedded within the protein-coding sequences that instructs the ribosome to ensure reading frame fidelity during translation.

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