scholarly journals Physiological Effects of Anti-TRAP Protein Activity and tRNATrp Charging on trp Operon Expression in Bacillus subtilis

2008 ◽  
Vol 190 (6) ◽  
pp. 1937-1945 ◽  
Author(s):  
Luis R. Cruz-Vera ◽  
Ming Gong ◽  
Charles Yanofsky
Keyword(s):  
Bacillus Subtilis ◽  
Growth Rates ◽  
Transcription Termination ◽  
Physiological Responses ◽  
Physiological Effects ◽  
Deficient Mutant ◽  
Wild Type ◽  
Protein Activity ◽  
Mutant Strains ◽  
Operon Expression

ABSTRACT The Bacillus subtilis anti-TRAP protein regulates the ability of the tryptophan-activated TRAP protein to bind to trp operon leader RNA and promote transcription termination. AT synthesis is regulated both transcriptionally and translationally by uncharged tRNATrp. In this study, we examined the roles of AT synthesis and tRNATrp charging in mediating physiological responses to tryptophan starvation. Adding excess phenylalanine to wild-type cultures reduced the charged tRNATrp level from 80% to 40%; the charged level decreased further, to 25%, in an AT-deficient mutant. Adding tryptophan with phenylalanine increased the charged tRNATrp level, implying that phenylalanine, when added alone, reduces the availability of tryptophan for tRNATrp charging. Changes in the charged tRNATrp level observed during growth with added phenylalanine were associated with increased transcription of the genes of tryptophan metabolism. Nutritional shift experiments, from a medium containing tryptophan to a medium with phenylalanine and tyrosine, showed that wild-type cultures gradually reduced their charged tRNATrp level. When this shift was performed with an AT-deficient mutant, the charged tRNATrp level decreased even further. Growth rates for wild-type and mutant strains deficient in AT or TRAP or that overproduce AT were compared in various media. A lack of TRAP or overproduction of AT resulted in phenylalanine being required for growth. These findings reveal the importance of AT in maintaining a balance between the synthesis of tryptophan versus the synthesis of phenylalanine, with the level of charged tRNATrp acting as the crucial signal regulating AT production.

scholarly journals Adenosine kinase-deficient mutant of Neurospora crassa

1982 ◽  
Vol 152 (3) ◽  
pp. 1292-1294
Author(s):  
J M Magill ◽  
P Dalke ◽  
T S Lyda ◽  
C W Magill
Keyword(s):  
Neurospora Crassa ◽  
Resistant Mutant ◽  
Adenosine Kinase ◽  
Deficient Mutant ◽  
Wild Type ◽  
Purine Nucleotides ◽  
Mutant Strains

Tubercidin-resistant mutant strains of Neurospora crassa were isolated, and at least one appeared to be deficient in adenosine kinase. No significant differences in [8-14C]adenosine labeling of purine nucleotides or nucleosides were found between the wild type and the adenosine kinase-deficient strains.

scholarly journals Transformation-deficient mutants of Bacillus subtilis impaired in competence-specific nuclease activities

1982 ◽  
Vol 152 (1) ◽  
pp. 166-174
Author(s):  
J A Mulder ◽  
G Venema
Keyword(s):  
Molecular Weight ◽  
Bacillus Subtilis ◽  
Sodium Dodecyl Sulfate ◽  
Sodium Dodecyl ◽  
Magnesium Ions ◽  
Polyacrylamide Gels ◽  
Wild Type ◽  
Molecular Weights ◽  
Defective Mutant ◽  
Mutant Strains

A comparison of the nucleolytic activities in competent and physiologically low-competent wild-type cultures of Bacillus subtilis in DNA-containing sodium dodecyl sulfate-polyacrylamide gels revealed the existence of three competence-associated nuclease activities with apparent molecular weights of 13,000, 15,000, and 26,000. The three activities, which were dependent on manganese or magnesium ions, were specifically present in the competent fraction of a competent culture. The competence-associated nucleolytic activities of eight transformation-defective mutant strains were assayed, resulting in the following three classes of mutants: (i) four strains which, according to this assay, were not impaired in any of the nucleolytic activities mentioned above; (ii) one strain which was strongly impaired in the 13,000- and 26,000-molecular-weight activities, but showed a considerable level of the 15,000-molecular-weight activity; and (iii) three strains which were severely impaired in all three activities. The results indicated that the 26,000-molecular-weight activity was a dimer of the 13,000-molecular-weight activity and that this nuclease was involved in the entry of DNA.

scholarly journals Addition of Poly(A) and Heteropolymeric 3′ Ends in Bacillus subtilis Wild-Type and Polynucleotide Phosphorylase-Deficient Strains

2005 ◽  
Vol 187 (14) ◽  
pp. 4698-4706 ◽  
Author(s):  
Juan Campos-Guillén ◽  
Patricia Bralley ◽  
George H. Jones ◽  
David H. Bechhofer ◽  
Gabriela Olmedo-Alvarez
Keyword(s):  
Bacillus Subtilis ◽  
Bacterial Species ◽  
Synthetic Activity ◽  
Polynucleotide Phosphorylase ◽  
Wild Type ◽  
Mutant Strains ◽  
Mean Size ◽  
Identical Nucleotide ◽  
End Sequences ◽  
Polymerase I

ABSTRACT Polyadenylation plays a role in decay of some bacterial mRNAs, as well as in the quality control of stable RNA. In Escherichia coli, poly(A) polymerase I (PAP I) is the main polyadenylating enzyme, but the addition of 3′ tails also occurs in the absence of PAP I via the synthetic activity of polynucleotide phosphorylase (PNPase). The nature of 3′-tail addition in Bacillus subtilis, which lacks an identifiable PAP I homologue, was studied. Sizing of poly(A) sequences revealed a similar pattern in wild-type and PNPase-deficient strains. Sequencing of 152 cloned cDNAs, representing 3′-end sequences of nontranslated and translated RNAs, revealed modified ends mostly on incomplete transcripts, which are likely to be decay intermediates. The 3′-end additions consisted of either short poly(A) sequences or longer heteropolymeric ends with a mean size of about 40 nucleotides. Interestingly, multiple independent clones exhibited complex heteropolymeric ends of very similar but not identical nucleotide sequences. Similar polyadenylated and heteropolymeric ends were observed at 3′ ends of RNA isolated from wild-type and pnpA mutant strains. These data demonstrated that, unlike the case of some other bacterial species and chloroplasts, PNPase of Bacillus subtilis is not the major enzyme responsible for the addition of nucleotides to RNA 3′ ends.

scholarly journals Uncovering New Metabolic Capabilities of Bacillus subtilis Using Phenotype Profiling of Rifampin-Resistant rpoB Mutants

2007 ◽  
Vol 190 (3) ◽  
pp. 807-814 ◽  
Author(s):  
Amy E. Perkins ◽  
Wayne L. Nicholson
Keyword(s):  
Bacillus Subtilis ◽  
Rna Polymerase ◽  
Metabolic Profiling ◽  
Transcriptional Control ◽  
Β Subunit ◽  
Subunit Gene ◽  
Wild Type ◽  
Mutant Strains ◽  
Utilization Patterns ◽  
Flow Of Information

ABSTRACT RNA polymerase is a central macromolecular machine controlling the flow of information from genotype to phenotype, and insights into global transcriptional regulation can be gained by studying mutational perturbations in the enzyme. Mutations in the RNA polymerase β subunit gene rpoB causing resistance to rifampin (Rifr) in Bacillus subtilis were previously shown to lead to alterations in the expression of a number of global phenotypes known to be under transcriptional control, such as growth, competence for transformation, sporulation, and germination (H. Maughan, B. Galeano, and W. L. Nicholson, J. Bacteriol. 186:2481-2486, 2004). To better understand the global effects of rpoB mutations on metabolism, wild-type and 11 distinct congenic Rifr mutant strains of B. subtilis were tested for utilization of 95 substrates by use of Biolog GP2 MicroPlates. A number of alterations of substrate utilization patterns were observed in the Rifr mutants, including the utilization of novel substrates previously unknown in B. subtilis, such as gentiobiose, β-methyl-d-glucoside, and d-psicose. The results indicate that combining global metabolic profiling with mutations in RNA polymerase provides a system-wide approach for uncovering previously unknown metabolic capabilities and further understanding global transcriptional control circuitry in B. subtilis.

scholarly journals Yersinia enterocolitica Induces Apoptosis and Inhibits Surface Molecule Expression and Cytokine Production in Murine Dendritic Cells

2004 ◽  
Vol 72 (12) ◽  
pp. 7045-7054 ◽  
Author(s):  
Stella E. Erfurth ◽  
Sabine Gröbner ◽  
Uwe Kramer ◽  
Dani S. J. Gunst ◽  
Irena Soldanova ◽  
...  
Keyword(s):  
Cell Proliferation ◽  
T Cells ◽  
Dendritic Cells ◽  
T Cell ◽  
Yersinia Enterocolitica ◽  
T Cell Proliferation ◽  
Surface Molecule ◽  
Deficient Mutant ◽  
Wild Type ◽  
Mutant Strains

ABSTRACT Yersinia enterocolitica evades innate immunity by expression of a variety of pathogenicity factors. Therefore, adaptive immunity including CD4+ T cells plays an important role in defense against Y. enterocolitica. We investigated whether Y. enterocolitica might target dendritic cells (DC) involved in adaptive T-cell responses. For this purpose, murine DC were infected with Y. enterocolitica wild-type and mutant strains prior to incubation with ovalbumin (OVA) as antigen and 5-(6)-carboxyfluorescein diacetate N-succinimidyl ester-labeled OVA-specific T cells from DO11.10 mice. While T-cell proliferation was partially affected by infection of DC with plasmid-cured and YopP-deficient Yersinia mutant strains, no T-cell proliferation occurred after infection of DC with wild-type Y. enterocolitica. Infection of DC with Y. enterocolitica wild type resulted in decreased up-regulation of major histocompatibility complex class II, CD54 (intercellular adhesion molecule 1), CD 80, and CD86 expression. Experiments with plasmid-cured Y. enterocolitica or a YopP-deficient mutant strain revealed that YopP accounts for inhibition of surface molecule expression. Wild-type Y. enterocolitica suppressed the release of KC, tumor necrosis factor alpha, interleukin-10 (IL-10), and IL-12 by DC, while infection of DC with plasmid-cured Y. enterocolitica or with the YopP-deficient mutant resulted in the production of these cytokines. Moreover, infection with wild-type Y. enterocolitica induced apoptosis in DC mediated by YopP. Apoptosis occurred despite translocation of NF-κB to the nucleus, as demonstrated by electromobility shift assays. Together, these data demonstrate that Y. enterocolitica targets functions of murine DC that are required for T-cell activation. This might contribute to evasion of adaptive immune responses by Y. enterocolitica.

Growth and photosynthetic activities of wild-type and antenna-deficient mutant strains of Rhodobacter capsulatus

1991 ◽  
Vol 155 (3) ◽  
pp. 205-209 ◽  
Author(s):  
Augusto F. Garcia ◽  
Werner M�ntele ◽  
Nasser Gad'on ◽  
Monier H. Tadros ◽  
Gerhart Drews
Keyword(s):  
Rhodobacter Capsulatus ◽  
Deficient Mutant ◽  
Wild Type ◽  
Mutant Strains ◽  
Photosynthetic Activities

scholarly journals Regulation of the Bacillus subtilis Extracytoplasmic Function Protein σY and Its Target Promoters

2003 ◽  
Vol 185 (16) ◽  
pp. 4883-4890 ◽  
Author(s):  
Min Cao ◽  
Letal Salzberg ◽  
Ching Sung Tsai ◽  
Thorsten Mascher ◽  
Carla Bonilla ◽  
...  
Keyword(s):  
Bacillus Subtilis ◽  
Sigma Factor ◽  
Negative Regulation ◽  
Wild Type ◽  
Induced Mutations ◽  
Mutant Strains ◽  
Elevated Expression ◽  
Direct Target ◽  
Extracytoplasmic Function Sigma Factor ◽  
Target Promoters

ABSTRACT The Bacillus subtilis extracytoplasmic function sigma factor σY is of unknown function. We demonstrate that the sigY operon is expressed from an autoregulatory promoter site, PY. We selected for transposon-induced mutations that upregulate PY transcription in an attempt to identify genes involved in σY regulation. The resulting insertions disrupted yxlC, the gene immediately downstream of sigY. However, the phenotype of the yxlC::Tn10 insertion was due to polarity on the downstream genes of the sigY operon; a nonpolar insertion in yxlC did not lead to derepression of PY. Further analyses revealed that both yxlD and yxlE encoded proteins important for the negative regulation of σY activity. A comparison of the transcriptomes of wild-type and yxlC::Tn10 mutant strains revealed elevated expression of several operons. However, only one additional gene, ybgB, was unambiguously identified as a direct target for σY. This was supported by analysis of direct targets for σY transcription with whole-genome runoff transcription followed by macroarray analysis.

scholarly journals Role of anionic phospholipids in the adaptation of Bacillus subtilis to high salinity

Microbiology ◽  
2006 ◽  
Vol 152 (3) ◽  
pp. 605-616 ◽  
Author(s):  
Claudia S. López ◽  
Alejandro F. Alice ◽  
Horacio Heras ◽  
Emilio A. Rivas ◽  
Carmen Sánchez-Rivas
Keyword(s):  
Bacillus Subtilis ◽  
Stationary Phase ◽  
Growth Curves ◽  
Membrane Lipid ◽  
Wild Type ◽  
Lipid Packing ◽  
Anionic Phospholipids ◽  
Large Unilamellar Vesicles ◽  
Mutant Strains ◽  
Lag Period

The importance of the content of anionic phospholipids [cardiolipin (CL) and phosphatidylglycerol (PG)] in the osmotic adaptation and in the membrane structure of Bacillus subtilis cultures was investigated. Insertion mutations in the three putative cardiolipin synthase genes (ywiE, ywnE and ywjE) were obtained. Only the ywnE mutation resulted in a complete deficiency in cardiolipin and thus corresponds to a true clsA gene. The osmotolerance of a clsA mutant was impaired: although at NaCl concentrations lower than 1·2 M the growth curves were similar to those of its wild-type control, at 1·5 M NaCl (LBN medium) the lag period increased and the maximal optical density reached was lower. The membrane of the clsA mutant strain showed an increased PG content, at both exponential and stationary phase, but no trace of CL in either LB or LBN medium. As well as the deficiency in CL synthesis, the clsA mutant showed other differences in lipid and fatty acids content compared to the wild-type, suggesting a cross-regulation in membrane lipid pathways, crucial for the maintenance of membrane functionality and integrity. The biophysical characteristics of membranes and large unilamellar vesicles from the wild-type and clsA mutant strains were studied by Laurdan's steady-state fluorescence spectroscopy. At physiological temperature, the clsA mutant showed a decreased lateral lipid packing in the protein-free vesicles and isolated membranes compared with the wild-type strain. Interestingly, the lateral lipid packing of the membranes of both the wild-type and clsA mutant strains increased when they were grown in LBN. In a conditional IPTG-controlled pgsA mutant, unable to synthesize PG and CL in the absence of IPTG, the osmoresistance of the cultures correlated with their content of anionic phospholipids. The transcriptional activity of the clsA and pgsA genes was similar and increased twofold upon entry to stationary phase or under osmotic upshift. Overall, these results support the involvement of the anionic phospholipids in the growth of B. subtilis in media containing elevated NaCl concentrations.

scholarly journals Influence of Bacillus subtilis strain on abscisic acid content in ABA-deficient barley mutant and its wild type

2021 ◽  
Vol 2(26) ◽  
pp. 28-40
Author(s):  
Z.A. Akhtyamova ◽  
◽  
T.N. Arkhipova ◽  
E.V. Martynenko ◽  
T.V. Nuzhnaya ◽  
...  
Keyword(s):  
Bacillus Subtilis ◽  
Abscisic Acid ◽  
Leaf Area ◽  
Photochemical Quenching ◽  
Bacterial Suspension ◽  
Subtilis Strain ◽  
Deficient Mutant ◽  
Wild Type ◽  
Promoting Effect ◽  
Shoot Mass

The ability to produce phytohormones and influence their metabolism in plants is an important property of rhizosphere bacteria that determines their plant growth promoting effect. Since abscisic acid (ABA) reduces stomatal conductance and increases the ability of tissues to conduct water, maintenance of water balance in lettuce plants on the background of activation of their growth was associated with the accumulation of ABA under the influence of bacteria. The aim of the study is to test the hypothesis that the growth-stimulating effect of bacteria on plants depends on their ability to synthesize the hormone ABA. The plants were grown on a light platform; seedlings were treated with a bacterial suspension simultaneously with planting. The ABA content, the relative water content, the chlorophyll content, the level of non-photochemical quenching, the leaf area and the weight of the shoots were measured. The level of transcripts of the HvNCED1, HvNCED2, and HvCYP707A1 genes responsible for ABA metabolism in barley was assessed using real-time PCR. Comparison of the ABA-deficient mutant of barley and plants of its wild type revealed the stimulation of the growth of plants of both genotypes upon bacterial treatment. The shoot mass and leaf area of the untreated mutant with bacteria were about 30 % less compared to Steptoe. The stimulating effect of bacteria was manifested in an increase in leaf area by 15 % in Steptoe and by 35 % in Az 34; shoot mass – by 18 % and 41 %, respectively. As a result, the phenotype difference between plants of two genotypes decreased. In the deficient mutant, the ABA level increased under the influence of Bacillus subtilis IB-22 more than twice. It was due to the ability of bacteria to produce ABA and reduce the activity of ABA degradation in barley plants. The results obtained in this study indicate that certain bacterial strains are able to increase the level of ABA in plants, compensating for the genetically determined deficiency of this hormone.

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