Knockout Mutation

2017 ◽  
Author(s):  
John H. Duffus ◽  
Michael Schwenk ◽  
Douglas M. Templeton
Keyword(s):  
Knockout Mutation

scholarly journals Deletion of znuA Virulence Factor Attenuates Brucella abortus and Confers Protection against Wild-Type Challenge

2006 ◽  
Vol 74 (7) ◽  
pp. 3874-3879 ◽  
Author(s):  
Xinghong Yang ◽  
Todd Becker ◽  
Nancy Walters ◽  
David W. Pascual
Keyword(s):  
Brucella Abortus ◽  
Deletion Mutant ◽  
Pasteurella Multocida ◽  
Brucella Melitensis ◽  
Normal Growth ◽  
Wild Type ◽  
Minimal Growth ◽  
M Genome ◽  
S19 Vaccine ◽  
Knockout Mutation

ABSTRACT znuA is known to be an important factor for survival and normal growth under low Zn2+ concentrations for Escherichia coli, Haemophilus spp., Neisseria gonorrhoeae, and Pasteurella multocida. We hypothesized that the znuA gene present in Brucella melitensis 16 M would be similar to znuA in B. abortus and questioned whether it may also be an important factor for growth and virulence of Brucella abortus. Using the B. melitensis 16 M genome sequence, primers were designed to construct a B. abortus deletion mutant. A znuA knockout mutation in B. abortus 2308 (ΔznuA) was constructed and found to be lethal in low-Zn2+ medium. When used to infect macrophages, ΔznuA B. abortus showed minimal growth. Further study with ΔznuA B. abortus showed that its virulence in BALB/c mice was attenuated, and most of the bacteria were cleared from the spleen within 8 weeks. Protection studies confirmed the ΔznuA mutant as a potential live vaccine, since protection against wild-type B. abortus 2308 challenge was as effective as that obtained with the RB51 or S19 vaccine strain.

scholarly journals Differential Gene Expression in Response to Hydrogen Peroxide and the Putative PerR Regulon of Synechocystis sp. Strain PCC 6803

2004 ◽  
Vol 186 (11) ◽  
pp. 3331-3345 ◽  
Author(s):  
Hong Li ◽  
Abhay K. Singh ◽  
Lauren M. McIntyre ◽  
Louis A. Sherman
Keyword(s):  
Chlorophyll Biosynthesis ◽  
Short Term ◽  
Histidine Kinases ◽  
Highly Sensitive ◽  
Number Of Genes ◽  
Genes Encoding ◽  
Differential Gene ◽  
Regulatory Properties ◽  
Knockout Mutation ◽  
Pcc 6803

ABSTRACT We utilized a full genome cDNA microarray to identify the genes that comprise the peroxide stimulon in the cyanobacterium Synechocystis sp. strain PCC 6803. We determined that a gene (slr1738) encoding a protein similar to PerR in Bacillus subtilis was induced by peroxide. We constructed a PerR knockout strain and used it to help identify components of the PerR regulon, and we found that the regulatory properties were consistent with the hypothesis that PerR functions as a repressor. This effort was guided by finding putative PerR boxes in positions upstream of specific genes and by careful statistical analysis. PerR and sll1621 (ahpC), which codes for a peroxiredoxin, share a divergent promoter that is regulated by PerR. We found that isiA, encoding a Chl protein that is induced under low-iron conditions, was strongly induced by a short-term peroxide stress. Other genes that were strongly induced by peroxide included sigD, sigB, and genes encoding peroxiredoxins and Dsb-like proteins that have not been studied yet in this strain. A gene (slr1894) that encoded a protein similar to MrgA in B. subtilis was upregulated by peroxide, and a strain containing an mrgA knockout mutation was highly sensitive to peroxide. A number of genes were downregulated, including key genes in the chlorophyll biosynthesis pathway and numerous regulatory genes, including those encoding histidine kinases. We used PerR mutants and a thioredoxin mutant (TrxA1) to study differential expression in response to peroxide and determined that neither PerR nor TrxA1 is essential for the peroxide protective response.

scholarly journals Ontogenetic Changes in Auxin Biosynthesis and Distribution Determine the Organogenic Activity of the Shoot Apical Meristem in pin1 Mutants

2019 ◽  
Vol 20 (1) ◽  
pp. 180 ◽  
Author(s):  
Alicja Banasiak ◽  
Magdalena Biedroń ◽  
Alicja Dolzblasz ◽  
Mateusz Adam Berezowski
Keyword(s):  
Shoot Apical Meristem ◽  
Apical Meristem ◽  
Auxin Transport ◽  
Vascular System ◽  
Auxin Biosynthesis ◽  
Wild Type ◽  
Ontogenetic Changes ◽  
Auxin Distribution ◽  
Stem Development ◽  
Knockout Mutation

In the shoot apical meristem (SAM) of Arabidopsis, PIN1-dependent polar auxin transport (PAT) regulates two crucial developmental processes: organogenesis and vascular system formation. However, the knockout mutation in the PIN1 gene does not fully inhibit these two processes. Therefore, we investigated a potential source of auxin for organogenesis and vascularization during inflorescence stem development. We analyzed auxin distribution in wild-type (WT) and pin1 mutant plants using a refined protocol of auxin immunolocalization; auxin activity, with the response reporter pDR5:GFP; and expression of auxin biosynthesis genes YUC1 and YUC4. Our results revealed that regardless of the functionality of PIN1-mediated PAT, auxin is present in the SAM and vascular strands. In WT plants, auxin always accumulates in all cells of the SAM, whereas in pin1 mutants, its localization within the SAM changes ontogenetically and is related to changes in the structure of the vascular system, organogenic activity of SAM, and expression levels of YUC1 and YUC4 genes. Our findings indicate that the presence of auxin in the meristem of pin1 mutants is an outcome of at least two PIN1-independent mechanisms: acropetal auxin transport from differentiated tissues with the use of vascular strands and auxin biosynthesis within the SAM.

scholarly journals Identification of a member of a DNA-dependent ATPase family that causes interference with silencing.

1997 ◽  
Vol 17 (9) ◽  
pp. 5461-5472 ◽  
Author(s):  
Z Zhang ◽  
A R Buchman
Keyword(s):  
Mating Type ◽  
Tandem Repeats ◽  
Specific Interaction ◽  
Cis Acting ◽  
Cellular Processes ◽  
Protein Dna Interactions ◽  
Mating Type Switching ◽  
Silencer Elements ◽  
Dna Template ◽  
Knockout Mutation

DNA in eukaryotic cells is packed in tandem repeats of nucleosomes or higher-order chromatin structures, which present obstacles to many cellular processes that require protein-DNA interactions, such as transcription, DNA repair, and recombination. To find proteins that are involved in increasing the accessibility of specific DNA regions in yeast, we used a genetic approach that exploited transcriptional silencing normally occurring at HML and HMR loci. The silencing is mediated by cis-acting silencer elements and is thought to require the formation of a special chromatin structure that prevents accessibility to the silenced DNA. A previously uncharacterized gene, termed DIS1, was isolated from a screen for genes that interfere with silencing when overexpressed. DIS1 encodes a protein with conserved motifs that are present in a family of DNA-dependent ATPases, the SWI2/SNF2-like proteins. Overproduction of N-terminal half of DIS1 protein interfered specifically with ectopic silencing used in the screen as well as HMR E silencing. Two-hybrid studies revealed a specific interaction between the N terminus of DIS1 and the C-terminal half of SIR4, a protein essential for silencing. Cells with a dis1 knockout mutation had significantly lower mating-type switching rate. These results suggest that DIS1 may contribute to making the silenced DNA template at HM loci more accessible during the mating-type switching process.

scholarly journals HRT-Mediated Hypersensitive Response and Resistance to Turnip crinkle virus in Arabidopsis Does Not Require the Function of TIP, the Presumed Guardee Protein

2008 ◽  
Vol 21 (10) ◽  
pp. 1316-1324 ◽  
Author(s):  
Rae-Dong Jeong ◽  
A. C. Chandra-Shekara ◽  
Aardra Kachroo ◽  
Daniel F. Klessig ◽  
Pradeep Kachroo
Keyword(s):  
Salicylic Acid ◽  
Nuclear Localization ◽  
Hypersensitive Response ◽  
Turnip Crinkle Virus ◽  
Basal Resistance ◽  
Interacting Protein ◽  
Enhanced Resistance ◽  
Resistance Protein ◽  
Knockout Mutation ◽  
Nac Family

The Arabidopsis resistance protein HRT recognizes the Turnip crinkle virus (TCV) coat protein (CP) to induce a hypersensitive response (HR) in the resistant ecotype Di-17. The CP also interacts with a nuclear-targeted NAC family of host transcription factors, designated TIP (TCV-interacting protein). Because binding of CP to TIP prevents nuclear localization of TIP, it has been proposed that TIP serves as a guardee for HRT. Here, we have tested the requirement for TIP in HRT-mediated HR and resistance by analyzing plants carrying knockout mutation in the TIP gene. Our results show that loss of TIP does not alter HR or resistance to TCV. Furthermore, the mutation in TIP neither impaired the salicylic acid–mediated induction of HRT expression nor the enhanced resistance conferred by overexpression of HRT. Strikingly, the mutation in TIP resulted in increased replication of TCV and Cucumber mosaic virus, suggesting that TIP may play a role in basal resistance but is not required for HRT-mediated signaling.

scholarly journals The ArgP Protein Stimulates the Klebsiella pneumoniae gdhA Promoter in a Lysine-Sensitive Manner

2008 ◽  
Vol 190 (12) ◽  
pp. 4351-4359 ◽  
Author(s):  
Thomas J. Goss
Keyword(s):  
Klebsiella Pneumoniae ◽  
Upstream Region ◽  
Mobility Shift ◽  
Base Pairs ◽  
Dnase I Footprinting ◽  
Sensitive Factor ◽  
Electrophoretic Mobility Shift Assays ◽  
Knockout Mutation

ABSTRACT The lysine-sensitive factor that binds to the upstream region of the Klebsiella pneumoniae gdhA promoter and stimulates gdhA transcription during growth in minimal medium has been proposed to be the K. pneumoniae ArgP protein (M. R. Nandineni, R. S. Laishram, and J. Gowrishankar, J. Bacteriol. 186:6391-6399, 2004). A knockout mutation of the K. pneumoniae argP gene was generated and used to assess the roles of exogenous lysine and argP in the regulation of the gdhA promoter. Disruption of argP reduced the strength and the lysine-dependent regulation of the gdhA promoter. Electrophoretic mobility shift assays using crude extracts prepared from wild-type and argP-defective strains indicted the presence of an argP-dependent factor whose ability to bind the gdhA promoter was lysine sensitive. DNase I footprinting studies using purified K. pneumoniae ArgP protein indicated that ArgP bound the region that lies approximately 50 to 100 base pairs upstream of the gdhA transcription start site in a manner that was sensitive to the presence of lysine. Substitutions within the region bound by ArgP affected the binding of ArgP to the gdhA promoter region in vitro and the argP-dependent stimulation of the gdhA promoter in vivo. These observations suggest that elevated intracellular levels of lysine reduce the affinity of ArgP for its binding site at the gdhA promoter, preventing ArgP from binding to and stimulating transcription from the promoter in vivo.

scholarly journals The role of B cells in lpr/lpr-induced autoimmunity.

1994 ◽  
Vol 180 (4) ◽  
pp. 1295-1306 ◽  
Author(s):  
M J Shlomchik ◽  
M P Madaio ◽  
D Ni ◽  
M Trounstein ◽  
D Huszar
Keyword(s):  
T Cells ◽  
B Cells ◽  
Primary Role ◽  
Tissue Destruction ◽  
Systemic Autoimmunity ◽  
Knockout Mutation

The primary roles of T cells and B cells in the initiation of systemic autoimmunity are unclear. To investigate the role of B cells, we crossed the "Jh knockout" mutation onto the autoimmune lpr/lpr background. Animals homozygous for both traits were obtained. As expected, these animals lack B cells. These animals also show no signs of autoimmune kidney destruction nor vasculitis, in spite of carrying the lpr/lpr mutation. In contrast, lpr/lpr littermates that had B cells had severe nephritis and vasculitis, as well as autoantibodies. These results demonstrate a primary role for B cells and/or (auto)antibodies in initiating several types of autoimmune-mediated tissue destruction. The implications of this finding for models and therapy of autoimmunity are discussed.

scholarly journals Author response: Programmed knockout mutation of liver fluke granulin attenuates virulence of infection-induced hepatobiliary morbidity

2018 ◽  
Author(s):  
Patpicha Arunsan ◽  
Wannaporn Ittiprasert ◽  
Michael J Smout ◽  
Christina J Cochran ◽  
Victoria H Mann ◽  
...  
Keyword(s):  
Liver Fluke ◽  
Author Response ◽  
Knockout Mutation
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