scholarly journals Regulation of Metalloprotease Gene Expression in Vibrio vulnificus by a Vibrio harveyi LuxR Homologue

2001 ◽  
Vol 183 (4) ◽  
pp. 1369-1375 ◽  
Author(s):  
Chung-Ping Shao ◽  
Lien-I Hor
Keyword(s):  
Parent Strain ◽  
Vibrio Vulnificus ◽  
Vibrio Harveyi ◽  
Negative Regulation ◽  
Open Reading Frame ◽  
Protease Gene ◽  
Reading Frame ◽  
Allelic Exchange ◽  
Exchange Technique

ABSTRACT Expression of the Vibrio vulnificus metalloprotease gene, vvp, was turned up rapidly when bacterial growth reached the late log phase. A similar pattern of expression has been found in the metalloprotease gene of Vibrio cholerae, and this has been shown to be regulated by a Vibrio harveyiLuxR-like transcriptional activator. To find out whether a LuxR homologue exists in V. vulnificus, a gene library of this organism was screened by colony hybridization using a probe derived from a sequence that is conserved in various luxR-like genes of vibrios. A gene containing a 618-bp open reading frame was identified and found to be identical to the smcR gene ofV. vulnificus reported previously. An isogenic SmcR-deficient (RD) mutant was further constructed by an in vivo allelic exchange technique. This mutant exhibited an extremely low level of vvp transcription compared with that of the parent strain. On the other hand, the cytolysin gene, vvhA, was expressed at a higher level in the RD mutant than in the parent strain during the log phase of growth. These data suggested that SmcR might not only be a positive regulator of the protease gene but might also be involved in negative regulation of the cytolysin gene. Virulence of the RD mutant in either normal or iron-overloaded mice challenged by intraperitoneal injection was comparable to that of the parent strain, indicating that SmcR is not required for V. vulnificusvirulence in mice.

scholarly journals vimA Gene Downstream of recAIs Involved in Virulence Modulation in Porphyromonas gingivalis W83

2001 ◽  
Vol 69 (1) ◽  
pp. 325-335 ◽  
Author(s):  
Hafid Abaibou ◽  
Zhuo Chen ◽  
G. Jon Olango ◽  
Yi Liu ◽  
Jessica Edwards ◽  
...  
Keyword(s):  
Porphyromonas Gingivalis ◽  
Parent Strain ◽  
Translational Regulation ◽  
Open Reading Frame ◽  
Transcriptional Unit ◽  
Reading Frame ◽  
Reverse Transcription Pcr ◽  
In Vivo Experiments ◽  
In The Wild

ABSTRACT A 0.9-kb open reading frame encoding a unique 32-kDa protein was identified downstream of the recA gene ofPorphyromonas gingivalis. Reverse transcription-PCR and Northern blot analysis showed that both the recA gene and this open reading frame are part of the same transcriptional unit. This cloned fragment was insertionally inactivated using theermF-ermAM antibiotic resistance cassette to create a defective mutant by allelic exchange. When plated onBrucella blood agar, the mutant strain, designated P. gingivalis FLL92, was non-black pigmented and showed significant reduction in beta-hemolysis compared with the parent strain,P. gingivalis W83. Arginine- and lysine-specific cysteine protease activities, which were mostly soluble, were approximately 90% lower than that of the parent strain. Expression of the rgpA, rgpB, and kgp protease genes was the same in P. gingivalis FLL92 as in the wild-type strain. In contrast to the parent strain, P. gingivalis FLL92 showed increased autoaggregration in addition to a significant reduction in hemagglutinating and hemolysin activities. In in vivo experiments using a mouse model, P. gingivalisFLL92 was dramatically less virulent than the parent strain. A molecular survey of this mutant and the parent strain using all knownP. gingivalis insertion sequence elements as probes suggested that no intragenomic changes due to the movement of these elements have occurred in P. gingivalis FLL92. Taken together, these results suggest that the recA downstream gene, designated vimA (virulence-modulating gene), plays an important role in virulence modulation in P. gingivalisW83, possibly representing a novel posttranscriptional or translational regulation of virulence factors in P. gingivalis.

scholarly journals Human Cytomegalovirus Open Reading Frame TRL11/IRL11 Encodes an Immunoglobulin G Fc-Binding Protein

2001 ◽  
Vol 75 (22) ◽  
pp. 11218-11221 ◽  
Author(s):  
Brendan N. Lilley ◽  
Hidde L. Ploegh ◽  
Rebecca S. Tirabassi
Keyword(s):  
Human Cytomegalovirus ◽  
Immunoglobulin G ◽  
Binding Protein ◽  
Fc Receptors ◽  
Binding Activity ◽  
Open Reading Frame ◽  
Membrane Glycoprotein ◽  
Type I ◽  
Reading Frame

ABSTRACT Several herpesviruses encode Fc receptors that may play a role in preventing antibody-mediated clearance of the virus in vivo. Human cytomegalovirus (HCMV) induces an Fc-binding activity in cells upon infection, but the gene that encodes this Fc-binding protein has not been identified. Here, we demonstrate that the HCMV AD169 open reading frame TRL11 and its identical copy, IRL11, encode a type I membrane glycoprotein that possesses IgG Fc-binding capabilities.

scholarly journals The Largest Open Reading Frame (pks12) in the Mycobacterium tuberculosis Genome Is Involved in Pathogenesis and Dimycocerosyl Phthiocerol Synthesis

2003 ◽  
Vol 71 (7) ◽  
pp. 3794-3801 ◽  
Author(s):  
Tatiana D. Sirakova ◽  
Vinod S. Dubey ◽  
Hwa-Jung Kim ◽  
Michael H. Cynamon ◽  
Pappachan E. Kolattukudy
Keyword(s):  
Fatty Acids ◽  
Mycobacterium Tuberculosis ◽  
Polyacrylamide Gel Electrophoresis ◽  
Sodium Dodecyl ◽  
Immunoblot Analysis ◽  
Open Reading Frame ◽  
Pcr Analysis ◽  
Reading Frame ◽  
Sds Page

ABSTRACT The cell wall lipids in Mycobacterium tuberculosis are probably involved in pathogenesis. The largest open reading frame in the genome of M. tuberculosis H37Rv, pks12, is unique in that it encodes two sets of domains needed to produce fatty acids. A pks12-disrupted mutant was produced, and disruption was confirmed by both PCR analysis and Southern blotting. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) analysis showed that a 430-kDa protein band present in the wild type was missing in the mutant. Matrix-assisted laser desorption ionization-time of flight mass spectrometry (MS) and liquid chromatography (LC)-MS analysis of tryptic peptides showed that 54 peptides distributed throughout this protein matched the pks12-encoded sequence. Biochemical analysis using [1-14C]propionate as the radiotracer showed that the pks12 mutant was deficient in the synthesis of dimycocerosyl phthiocerol (DIM). SDS-PAGE, immunoblot analysis of proteins, and analysis of fatty acids showed that the mutant can produce mycocerosic acids. Thus, the pks12 gene is probably involved in the synthesis of phthiocerol, the diol required for DIM synthesis. Growth of the pks12 mutant was attenuated in mouse alveolar macrophage cell line MH-S, and the virulence of the mutant in vivo was highly attenuated in a murine model. Thus, pks12 probably participates in DIM production and its expression is involved in pathogenesis.

scholarly journals Murine cytomegalovirus open reading frame m29.1 augments virus replication both in vitro and in vivo

2007 ◽  
Vol 88 (11) ◽  
pp. 2941-2951 ◽  
Author(s):  
Mohammad M. Ahasan ◽  
Clive Sweet
Keyword(s):  
Virus Replication ◽  
Post Infection ◽  
Stop Codon ◽  
Premature Stop Codon ◽  
Open Reading Frame ◽  
Murine Cytomegalovirus ◽  
Reading Frame ◽  
Codon Mutation

Murine cytomegalovirus mutant Rc29, with a premature stop codon mutation in the m29 open reading frame (ORF), produced no apparent phenotype in cell culture or following infection of BALB/c mice. In contrast, a similar mutant virus, Rc29.1, with a premature stop codon mutation in its m29.1 ORF, showed reduced virus yields (2–3 log10 p.f.u. ml−1) in tissue culture. Mutant virus yields in BALB/c mice were delayed, reduced (∼1 log10 p.f.u. per tissue) and persisted less well in salivary glands compared with wild-type (wt) and revertant (Rv29.1) virus. In severe combined immunodeficiency mice, Rc29.1 virus showed delayed and reduced replication initially in all tissues (liver, spleen, kidneys, heart, lung and salivary glands). This delayed death until 31 days post-infection (p.i.) compared with wt (23 days p.i.) but at death virus yields were similar to wt. m29 gene transcription was initiated at early times post-infection, while production of a transcript from ORF m29.1 in the presence of cycloheximide indicated that it was an immediate-early gene. ORFs m29.1 and M28 are expressed from a bicistronic message, which is spliced infrequently. However, it is likely that each ORF expresses its own protein, as antiserum derived in rabbits to the m29.1 protein expressed in bacteria from the m29.1 ORF detected only one protein in Western blot analysis of the size predicted for the m29.1 protein. Our results suggest that neither ORF is essential for virus replication but m29.1 is important for optimal viral growth in vitro and in vivo.

scholarly journals Generation of an Adenovirus Vector Lacking E1, E2a, E3, and All of E4 except Open Reading Frame 3

1999 ◽  
Vol 73 (7) ◽  
pp. 6048-6055 ◽  
Author(s):  
Mario I. Gorziglia ◽  
Claudia Lapcevich ◽  
Soumitra Roy ◽  
Qiang Kang ◽  
Mike Kadan ◽  
...  
Keyword(s):  
Gene Therapy ◽  
Liver Toxicity ◽  
Adenovirus Vector ◽  
Open Reading Frame ◽  
Reading Frame ◽  
Rous Sarcoma ◽  
Sarcoma Virus ◽  
Adenovirus Vectors ◽  
Virus Promoter

ABSTRACT Toxicity and immunity associated with adenovirus backbone gene expression is an important hurdle to overcome for successful gene therapy. Recent efforts to improve adenovirus vectors for in vivo use have focused on the sequential deletion of essential early genes. Adenovirus vectors have been constructed with the E1 gene deleted and with this deletion in combination with an E2a, E2b, or E4 deletion. We report here a novel vector (Av4orf3nBg) lacking E1, E2a, and all of E4 except open reading frame 3 (ORF3) and expressing a β-galactosidase reporter gene. This vector was generated by transfection of a plasmid carrying the full-length vector sequence into A30.S8 cells that express E1 and E2a but not E4. Production was subsequently performed in an E1-, E2a-, and E4-complementing cell line. We demonstrated with C57BL/6 mice that the Av4orf3nBg vector effected gene transfer with an efficiency comparable to that of the Av3nBg (wild-type E4) vector but that the former exhibited a higher level of β-galactosidase expression. This observation suggests that E4 ORF3 alone is able to enhance RNA levels from the β-galactosidase gene when the Rous sarcoma virus promoter is used to drive transgene expression in the mouse liver. In addition, we observed less liver toxicity in mice injected with the Av4orf3nBg vector than those injected with the Av3nBg vector at a comparable DNA copy number per cell. This study suggests that the additional deletion of E4 in an E1 and E2a deletion background may be beneficial in decreasing immunogenicity and improving safety and toxicity profiles, as well as increasing transgene capacity and expression for liver-directed gene therapy.

scholarly journals Neuronal Cell-type Engineering by Transcriptional Activation

2021 ◽  
Vol 3 ◽  
Author(s):  
Songlei Liu ◽  
Johannes Striebel ◽  
Giovanni Pasquini ◽  
Alex H. M. Ng ◽  
Parastoo Khoshakhlagh ◽  
...  
Keyword(s):  
Transcriptional Activation ◽  
Recent Progress ◽  
Gene Activation ◽  
Neuronal Cell ◽  
Open Reading Frame ◽  
Cell Type ◽  
Reading Frame ◽  
Cellular Behavior ◽  
Technical Parameters

Gene activation with the CRISPR-Cas system has great implications in studying gene function, controlling cellular behavior, and modulating disease progression. In this review, we survey recent studies on targeted gene activation and multiplexed screening for inducing neuronal differentiation using CRISPR-Cas transcriptional activation (CRISPRa) and open reading frame (ORF) expression. Critical technical parameters of CRISPRa and ORF-based strategies for neuronal programming are presented and discussed. In addition, recent progress on in vivo applications of CRISPRa to the nervous system are highlighted. Overall, CRISPRa represents a valuable addition to the experimental toolbox for neuronal cell-type programming.

scholarly journals Ilarviruses Encode a Cucumovirus-Like 2b Gene That Is Absent in Other Genera within the Bromoviridae

1998 ◽  
Vol 72 (8) ◽  
pp. 6956-6959 ◽  
Author(s):  
Hong-Wu Xin ◽  
Liang-Hui Ji ◽  
Simon W. Scott ◽  
Robert H. Symons ◽  
Shou-Wei Ding
Keyword(s):  
Open Reading Frame ◽  
Latent Virus ◽  
Evolutionary Origin ◽  
Protein Species ◽  
Subgenomic Rna ◽  
Reading Frame ◽  
2B Protein ◽  
Sequence Similarities

ABSTRACT We found that RNA 2 of the four ilarviruses sequenced to date encodes an additional conserved open reading frame (ORF), 2b, that overlaps the 3′ end of the previously known ORF, 2a. A novel RNA species of 851 nucleotides was found to accumulate to high levels in plants infected with spinach latent virus (SpLV). Further analysis showed that RNA 4A is a subgenomic RNA of RNA 2 and encodes all of ORF 2b. Moreover, a protein species of the size expected for SpLV ORF 2b was translated in vitro from the RNA 4A-containing virion RNAs. The data support the suggestion that the SpLV 2b protein is translated in vivo. The 2b gene of ilarviruses, which is not encoded by alfamoviruses and bromoviruses, shares several features with the previously reported cucumovirus 2b gene; however, their encoded proteins share no detectable sequence similarities. The evolutionary origin of the 2b gene is discussed.

scholarly journals Translation rescue by targeting Ppp1r15a upstream open reading frame in vivo

2021 ◽  
Author(s):  
Ashley Kidwell ◽  
Shiv Pratap Singh Yadav ◽  
Bernhard Maier ◽  
Amy Zollman ◽  
Kevin Ni ◽  
...  
Keyword(s):  
Kidney Injury ◽  
Open Reading Frame ◽  
Upstream Open Reading Frame ◽  
Regulatory Subunit ◽  
Reading Frame ◽  
Extreme Stress ◽  
Eif2α Phosphorylation ◽  
Endotoxemia Model ◽  
Life Threatening

The eIF2 initiation complex is central to maintaining a functional translation machinery. Extreme stress such as life-threatening sepsis exposes vulnerabilities in this tightly regulated system, resulting in an imbalance between the opposing actions of kinases and phosphatases on the main regulatory subunit eIF2α. Here, we report that translation shutdown is a hallmark of established sepsis-induced kidney injury brought about by excessive eIF2α phosphorylation and sustained by blunted expression of the counterregulatory phosphatase subunit Ppp1r15a. We determined that the blunted Ppp1r15a expression persists because of the presence of an upstream open reading frame (uORF). Overcoming this barrier with genetic approaches enabled the derepression of Ppp1r15a, salvaged translation and improved kidney function in an endotoxemia model. We also found that the loss of this uORF has broad effects on the composition and phosphorylation status of the immunopeptidome that extended beyond the eIF2α axis. Collectively, our findings define the breath and potency of the highly conserved Ppp1r15a uORF and provide a paradigm for the design of uORF-based translation rheostat strategies. The ability to accurately control the dynamics of translation during sepsis will open new paths for the development of therapies at codon level precision.

Molecular biology and pathogenesis of hepatitis E virus

1999 ◽  
Vol 1 (18) ◽  
pp. 1-16 ◽  
Author(s):  
Shahid Jameel
Keyword(s):  
Viral Genome ◽  
Hepatitis E Virus ◽  
Hepatitis E ◽  
Open Reading Frame ◽  
Reading Frame ◽  
Host Interactions ◽  
Processing Enzymes ◽  
Open Reading Frame 2

Hepatitis E virus (HEV) infection results in hepatitis E, an acute and self-limited disease. The virus is transmitted in a faecal–oral manner and is a major cause of viral hepatitis in much of the developing world, where it causes rampant sporadic infections and large epidemics. A curious feature of hepatitis E is the unusually high rates of mortality that are observed in pregnant women, in whom the disease is exacerbated by the development of fulminant liver disease. In the absence of viable in vitro propagation systems, several geographical isolates of HEV have been maintained in vivo in nonhuman primates and, subsequently, the viral genome has been cloned and sequenced. HEV has been classified provisionally into a separate family known as the HEV-like viruses, which has at least four recognised genotypes, but has only a single serotype. The viral genome is a positive-stranded (+)RNA of ~7.5 kb and encodes at least three proteins. Open reading frame 1 (ORF1) encodes the viral nonstructural polyprotein, which has domains that are homologous to some of the replication and processing enzymes found in other +RNA viruses. The HEV protein itself remains poorly characterised. The protein encoded by open reading frame 2 (ORF2) is the major HEV capsid protein, and the protein encoded by open reading frame 3 (ORF3) appears to be involved in virus–host interactions. Several questions related to the biology, epidemiology and pathogenesis of HEV remain unanswered; the progress of a few of these is reviewed here.

Sign in / Sign up

Export Citation Format

Share Document