Regulation of ribosomal protein synthesis in mycobacteria: autogenous control of rpsO

ABSTRACTAutogenous regulation of ribosomal protein (r-protein) synthesis plays a key role in maintaining the stoichiometry of ribosomal components in bacteria. Our main goal was to develop techniques for investigating the r-protein synthesis regulation in mycobacteria, Gram-positive organisms with a high GC-content, which has never been addressed. We started with the rpsO gene known to be autoregulated by its product, r-protein S15, in a broad range of bacterial species. To study the in vivo regulation of rpsO from Mycobacterium smegmatis (Msm), we first applied an approach based on chromosomally integrated Msm rpsO’-’lacZ reporters by using E. coli as a surrogate host. The β-galactosidase assay has shown that mycobacterial rpsO expression is feedback regulated at the translation level in the presence of Msm S15 in trans, like in E. coli. Next, to overcome difficulties caused by the inefficiency of mycobacterial gene expression in E. coli, we created a fluorescent reporter system based on M. smegmatis. To this end, the integrative shuttle plasmid pMV306 was modified to provide insertion of the Msm or Mtb (M. tuberculosis) rpsO-egfp reporters into the Msm chromosome, and a novel E. coli-mycobacteria replicative shuttle vector, pAMYC, a derivative of pACYC184, was built. Analysis of the eGFP expression in the presence of the pAMYC derivative expressing Msm rpsO vs an empty vector confirms the autogenous regulation of the rpsO gene in mycobacteria. Additionally, we have revealed that the mycobacterial rpsO core promoters are rather weak and require upstream activating elements to enhance their strength.IMPORTANCEBacterial ribosomes are targets for a majority of as-yet reported antibiotics, hence ribosome biogenesis and its regulation are central for development of new antimicrobials. One of the key mechanisms regulating ribosome biogenesis in bacteria is the autogenous control of r-protein synthesis, which has been so far explored for E. coli and Bacillus spp. but not yet for mycobacteria. Here, we describe experimental approaches for in vivo analysis of mechanisms regulating r-protein synthesis in mycobacteria, including M. tuberculosis, and show, for the first time, that the autogenous control at the translation level is really functioning in these microorganisms. The developed system paves the way for studying various regulatory circuits involving proteins or sRNAs as mRNA- targeting trans-regulators in mycobacteria as well as in other actinobacterial species.

Autogenous control of heart rate taking Deep Slow Breaths during exercise

We investigated whether heart rate can be controlled consciously. This study examined the effect of 20 minutes of cycling exercise while being conscious about energy conservation on the heart rate. 21 healthy college students (9 men, 12 women) participated three exercise bouts. Exercise bouts were examined under the following three conditions: (a) known condition (participants having information about the exercise duration); (b) unknown condition (participants having no information about the exercise duration) and; (c) conserving energy condition (participants having information about the exercise duration and being conscious about energy conservation). Heart rate in the unknown condition was lower than that in the known condition (p < 0.05); further, it was lower in the conserving energy condition than in the known condition (p < 0.01). In contrast, the tidal volume of the conserving energy condition was higher than that in the known condition (p < 0.05). In addition, the respiratory rates in the unknown and conserving energy conditions were lower than that in the known condition (p < 0.01). Energy expenditure during exercise was lowest in the conserving energy condition. These results show the possibility of autogenous control of heart rate by taking deep slow breaths consciously during exercise.

Autogenous control augmentation system – A refinement in diced cartilage glue graft for augmentation of dorsum of nose

ABSTRACT Background: In the context of different grafts being used for dorsal augmentation, diced cartilage with glue has gained worldwide acceptance. Aims: To develop a system of tools to objectively evaluate the desired dimensions of the required graft for dorsal augmentation and to prepare a corresponding customized-glued-diced cartilage construct. Materials and Methods: A modification of the diced cartilage glue technique called Autogenous control augmentation system (ACAS) was used in ten patients. Results: Of the ten patients, in which this technique was used, eight underwent primary rhinoplasties and two underwent secondary rhinoplasties between July 2017 and December 2017 with a follow-up ranging from 3 to 8 months. In all the cases, the dorsum is straight, and height is maintained. Conclusion: The technique has all the advantages of the diced cartilage glue. The shape resembles alloplastic implant with height and width varying from radix to tip. There is tapering of the cephalic and caudal ends for more natural results. The brow tip aesthetic lines are better defined. The limitation of this study is short follow up.

Negative Autogenous Control of the Master Type III Secretion System Regulator HrpL in Pseudomonas syringae

ABSTRACT The type III secretion system (T3SS) is a principal virulence determinant of the model bacterial plant pathogen Pseudomonas syringae. T3SS effector proteins inhibit plant defense signaling pathways in susceptible hosts and elicit evolved immunity in resistant plants. The extracytoplasmic function sigma factor HrpL coordinates the expression of most T3SS genes. Transcription of hrpL is dependent on sigma-54 and the codependent enhancer binding proteins HrpR and HrpS for hrpL promoter activation. hrpL is oriented adjacently to and divergently from the HrpL-dependent gene hrpJ, sharing an intergenic upstream regulatory region. We show that association of the RNA polymerase (RNAP)-HrpL complex with the hrpJ promoter element imposes negative autogenous control on hrpL transcription in P. syringae pv. tomato DC3000. The hrpL promoter was upregulated in a ΔhrpL mutant and was repressed by plasmid-borne hrpL. In a minimal Escherichia coli background, the activity of HrpL was sufficient to achieve repression of reconstituted hrpL transcription. This repression was relieved if both the HrpL DNA-binding function and the hrp-box sequence of the hrpJ promoter were compromised, implying dependence upon the hrpJ promoter. DNA-bound RNAP-HrpL entirely occluded the HrpRS and partially occluded the integration host factor (IHF) recognition elements of the hrpL promoter in vitro, implicating inhibition of DNA binding by these factors as a cause of negative autogenous control. A modest increase in the HrpL concentration caused hypersecretion of the HrpA1 pilus protein but intracellular accumulation of later T3SS substrates. We argue that negative feedback on HrpL activity fine-tunes expression of the T3SS regulon to minimize the elicitation of plant defenses. IMPORTANCE The United Nations Food and Agriculture Organization has warned that agriculture will need to satisfy a 50% to 70% increase in global food demand if the human population reaches 9 billion by 2050 as predicted. However, diseases caused by microbial pathogens represent a major threat to food security, accounting for over 10% of estimated yield losses in staple wheat, rice, and maize crops. Understanding the decision-making strategies employed by pathogens to coordinate virulence and to evade plant defenses is vital for informing crop resistance traits and management strategies. Many plant-pathogenic bacteria utilize the needle-like T3SS to inject virulence factors into host plant cells to suppress defense signaling. Pseudomonas syringae is an economically and environmentally devastating plant pathogen. We propose that the master regulator of its entire T3SS gene set, HrpL, downregulates its own expression to minimize elicitation of plant defenses. Revealing such conserved regulatory strategies will inform future antivirulence strategies targeting plant pathogens. The United Nations Food and Agriculture Organization has warned that agriculture will need to satisfy a 50% to 70% increase in global food demand if the human population reaches 9 billion by 2050 as predicted. However, diseases caused by microbial pathogens represent a major threat to food security, accounting for over 10% of estimated yield losses in staple wheat, rice, and maize crops. Understanding the decision-making strategies employed by pathogens to coordinate virulence and to evade plant defenses is vital for informing crop resistance traits and management strategies. Many plant-pathogenic bacteria utilize the needle-like T3SS to inject virulence factors into host plant cells to suppress defense signaling. Pseudomonas syringae is an economically and environmentally devastating plant pathogen. We propose that the master regulator of its entire T3SS gene set, HrpL, downregulates its own expression to minimize elicitation of plant defenses. Revealing such conserved regulatory strategies will inform future antivirulence strategies targeting plant pathogens.

Dual autogenous control of the multiple antibiotic resistance phenotype in Escherichia coli

Bacteria can defend against diverse antibiotics by mounting a multiple antibiotic resistance (mar) phenotype. The resistance is linked to a chromosomal locus that encodes an activator and a repressor regulating their own expression. Here, we investigated how this dual autogenous control determines the dynamics of the response. We found that the regulatory architecture provides a mechanism to enable rapid induction, generate pulses of activation, and increase the range of sensing. The response is also graded and homogeneous across the population. Moreover, the interaction of a third regulator with the core module fine tunes the previous features, while limiting the cross-talk with metabolic signals. A minimal model accurately anticipates these properties, and emphasizes how specific attributes of the circuit components constrain the appearance of other potential behaviors associated to the regulatory design. Our results integrate both molecular and circuit-level characteristics to fully elucidate the dynamic emergence of the mar phenotype.

Pipe Rupture Analysis Considering Fluid-Structure Interaction

Global warming is caused by the emission of greenhouse gases, like CO2. Nuclear energy is one of the main sources of low-carbon energy. In the events of serious accidents, a nuclear power plant may emit radioactivity that is harmful to human health. Nuclear power should be used after enough evidence of its safety is provided. Measures for safety of nuclear power plants, such as autogenous control and LBB, have been developed. Moreover, there is requirement with respect to the design, safety, equipments components and systems of nuclear plant. For example, it is necessary to place components that restrain pipe whip behavior, and to design peripheral equipments that may be affected by high-pressured fluid in pipe rupture accidents [1], [2]. In the case of pipe rupture that occurs to structures such as nuclear plants and steam generators, a pipe deforms releasing its inner high-pressured fluid. In previous studies, the pipe whip behavior analyses have been performed by using blowdown thrust force that is estimated by fluid analysis. In this study, we simulate pipe whip behavior and reduction of blowdown thrust force by releasing inner fluid to the atmosphere. The analysis model is an elbow pipe and high-pressure fluid running inside. We considered fluid-structure interaction effect in the analysis because ovalization of the cross-section of the elbow part as well as a change of the elbow torus radius affects fluid flow blowing out from the ruptured part of the pipe.