scholarly journals Cellular NADH and NADPH Conformation as a Real-Time Fluorescence-Based Metabolic Indicator under Pressurized Conditions

Molecules ◽  
2021 ◽  
Vol 26 (16) ◽  
pp. 5020
Author(s):  
Martin Heidelman ◽  
Bibek Dhakal ◽  
Millicent Gikunda ◽  
Kalinga Pavan Thushara Silva ◽  
Laxmi Risal ◽  
...  

Cellular conformation of reduced pyridine nucleotides NADH and NADPH sensed using autofluorescence spectroscopy is presented as a real-time metabolic indicator under pressurized conditions. The approach provides information on the role of pressure in energy metabolism and antioxidant defense with applications in agriculture and food technologies. Here, we use spectral phasor analysis on UV-excited autofluorescence from Saccharomyces cerevisiae (baker’s yeast) to assess the involvement of one or multiple NADH- or NADPH-linked pathways based on the presence of two-component spectral behavior during a metabolic response. To demonstrate metabolic monitoring under pressure, we first present the autofluorescence response to cyanide (a respiratory inhibitor) at 32 MPa. Although ambient and high-pressure responses remain similar, pressure itself also induces a response that is consistent with a change in cellular redox state and ROS production. Next, as an example of an autofluorescence response altered by pressurization, we investigate the response to ethanol at ambient, 12 MPa, and 30 MPa pressure. Ethanol (another respiratory inhibitor) and cyanide induce similar responses at ambient pressure. The onset of non-two-component spectral behavior upon pressurization suggests a change in the mechanism of ethanol action. Overall, results point to new avenues of investigation in piezophysiology by providing a way of visualizing metabolism and mitochondrial function under pressurized conditions.

2019 ◽  
Vol 116 (3) ◽  
pp. 567a-568a
Author(s):  
Paul K. Urayama ◽  
Audrey Short ◽  
Martin Heidelman ◽  
Max Kreider ◽  
Andrew I. Rodriguez ◽  
...  

2008 ◽  
Vol 190 (13) ◽  
pp. 4624-4631 ◽  
Author(s):  
Hideaki Mizusaki ◽  
Akiko Takaya ◽  
Tomoko Yamamoto ◽  
Shin-Ichi Aizawa

ABSTRACT Salmonella enterica serovar Typhimurium secretes virulence factors for invasion called Sip proteins or Sips into its hosts through a type III secretion system (T3SS). In the absence of a host, S. enterica induces Sip secretion in response to sucrose or simple salts, such as NaCl. We analyzed induction of host-independent Sip secretion by monitoring protein secretion by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE), assembly of needle complexes by electron microscopy, and transcription of virulence regulatory genes by quantitative reverse transcriptase PCR (real-time PCR). SDS-PAGE showed that addition of sucrose or simple salts, such as NaCl, to the growth medium induced Sip secretion without altering flagellar protein secretion, which requires a distinct T3SS. Electron microscopy confirmed that the amount of secreted Sips increased as the number of assembled needle complexes increased. Real-time PCR revealed that added sucrose or NaCl enhanced transcription of hilA, hilC, and hilD, which encode known regulators of Salmonella virulence. However, epistasis analysis implicated HilD and HilA, but not HilC, in the direct pathway from the salt stimulus to the Sip secretion response. Further analyses showed that the BarA/SirA two-component signal transduction pathway, but not the two-component sensor kinase EnvZ, directly activated hilD and hilA transcription and thus Sip secretion in response to either sucrose or NaCl. Finally, real-time PCR showed that salt does not influence transcription of the BarA/SirA-dependent csrB and csrC genes. A model is proposed for the major pathway in which sucrose or salt signals to enhance virulence gene expression.


2021 ◽  
Vol 21 (1) ◽  
Author(s):  
Sanaz Dehbashi ◽  
Hamed Tahmasebi ◽  
Behrouz Zeyni ◽  
Mohammad Reza Arabestani

Abstract Background Methicillin-resistant Staphylococcus aureus (MRSA)-bloodstream infections (BSI) are predominantly seen in the hospital or healthcare-associated host. Nevertheless, the interactions of virulence factor (VFs) regulators and β-lactam resistance in MRSA-BSI are unclear. This study aims to characterize the molecular relationship of two-component systems of VFs and the expression of the β-lactamase gene in MRSA-BSI isolates. In this study, 639 samples were collected from BSI and identified by phenotypic methods. We performed extensive molecular characterization, including SCCmec type, agr type, VFs gene profiles determinations, and MLST on isolates. Also, a quantitative real-time PCR (q-RT PCR) assay was developed for identifying the gene expressions. Results Ninety-one (91) S. aureus and 61 MRSA (67.0%) strains were detected in BSI samples. The presence of VFs and SCCmec genes in MRSA isolates were as follows: tst (31.4%), etA (18.0%), etB (8.19%), lukS-PVL (31.4%), lukF-PV (18.0%), lukE-lukD (16.3%), edin (3.2%), hla (16.3%), hlb (18.0%), hld (14.7%), hlg (22.9%), SCCmecI (16.3%), SCCmecII (22.9%), SCCmecIII (36.0%), SCCmecIV (21.3%), and SCCmecV (16.3%). Quantitative real-time PCR showed overexpression of mecRI and mecI in the toxigenic isolates. Moreover, RNAIII and sarA genes were the highest expressions of MRSA strains. The multi-locus sequence typing data confirmed a high prevalence of CC5, CC8, and CC30. However, ST30, ST22, and ST5 were the most prevalent in the resistant and toxigenic strains. Conclusion We demonstrated that although regulation of β-lactamase gene expressions is a significant contributor to resistance development, two-component systems also influence antibiotic resistance development in MRSA-BSI isolates. This indicates that resistant strains might have pathogenic potential. We also confirmed that some MLST types are more successful colonizers with a potential for MRSA-BSI.


2008 ◽  
Vol 92 (1) ◽  
pp. 012110 ◽  
Author(s):  
Yoshiharu Enta ◽  
Bongjin S. Mun ◽  
Massimiliano Rossi ◽  
Philip N. Ross ◽  
Zahid Hussain ◽  
...  

1998 ◽  
Author(s):  
Orven F. Swenson ◽  
Josef P. Carriere ◽  
Harlan Isensee ◽  
Gregory D. Gillispie ◽  
William F. Cooper ◽  
...  

2016 ◽  
Vol 145 (10) ◽  
pp. 104107 ◽  
Author(s):  
Joshua J. Goings ◽  
Joseph M. Kasper ◽  
Franco Egidi ◽  
Shichao Sun ◽  
Xiaosong Li

2018 ◽  
Vol 210 ◽  
pp. 429-449 ◽  
Author(s):  
A. Robert Hillman ◽  
Robert Barker ◽  
Robert M. Dalgliesh ◽  
Virginia C. Ferreira ◽  
Emma J. R. Palin ◽  
...  

We investigate the deposition, evolution and dissolution of single and two-component metal layers on Au substrates immersed in the deep eutectic solvent Ethaline.


2021 ◽  
Author(s):  
Janet Sorrells ◽  
Rishyashring Iyer ◽  
Lingxiao Yang ◽  
Andrew Bower ◽  
Darold Spillman ◽  
...  

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