Unraveling the Molecular Mechanisms Underlying the Microbiome Response to Soil Rewetting

Soil microbes are highly sensitive to changes in their environment, and rapidly measuring their responses is necessary to fully understand the biological processes. Drought is one of the most common environmental stresses that soil microbiomes experience, and it is important to understand the mechanisms by which the soil microbiome respond to soil dehydration. We used 13C as a tracer of nutrient fluxes in desiccated soil microbiomes after rewetting to simultaneously measure aerobic respiration and track the metabolic state of the community. Here, we describe a Real Time Mass Spectrometry (RTMS) approach for rapid gas monitoring combined with omics approaches to track 13C flow through a soil system.The mechanism(s) behind the burst of rapid mineralization of soil organic matter and increased rate of CO2&#160;release upon rewetting dry soil (termed the &#8216;Birch Effect&#8217;) are yet to be fully defined. One known mechanism used by microbes to protect against dehydration is the production of intracellular compounds known as osmolytes. We evaluated metabolic mechanisms produced upon rewetting a marginal soil testing the hypothesis that the rapid release of CO2&#160;arises from the microbial processing of putative intracellular osmolytes that build up during desiccation. RTMS allows for the simultaneous, rapid and fine scale (every 2 sec) evaluation and deconvolution of the production and consumption of a number of gasses including 12CO2,13CO2, O2, N2&#160;and H2O. &#160;We compared the hydration response (production of CO2&#160;in real time) between the addition of water and 13C labeled glucose dissolved in water. The initial burst of 12CO2&#160;followed by a leveling off was identical in both treatments with an additional larger increase in 13CO2&#160;about 20 minutes later in the 13C labeled glucose experiment. Examination of the two minutes after the water addition revealed a rapid rate of 12CO2 (38 sec) and H2O (47 sec) production and slow rate of 13CO2 (56 sec) production followed by the consumption of O2&#160;(67 sec) and N2 (73 sec).&#160; Evaluation of the soil metabolomes at specified time points within 3 hours after wetting revealed the immediate release of sugars from the cells into the extracellular matrix. These results provide evidence for respiration of putative intracellular osmolytes as one driving mechanism of the Birch Effect.&#160;

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RPS24c Isoform Facilitates Tumor Angiogenesis Via Promoting the Stability of MVIH in Colorectal Cancer

Current Molecular Medicine ◽

10.2174/1566524019666191203123943 ◽

2020 ◽

Vol 20 (5) ◽

pp. 388-395 ◽

Cited By ~ 1

Author(s):

Yue Wang ◽

Youjun Wu ◽

Kun Xiao ◽

Yingjie Zhao ◽

Gang Lv ◽

...

Keyword(s):

Colorectal Cancer ◽

Ribosomal Protein ◽

Real Time ◽

Tumor Angiogenesis ◽

Real Time Pcr ◽

Molecular Mechanisms ◽

Migration And Invasion ◽

Binding Interaction ◽

Tubule Formation ◽

The Stability

Background: Colorectal cancer (CRC) is the second leading cause of death worldwide, and distant metastasis is responsible for the poor prognosis in patients with advanced-stage CRC. RPS24 (ribosomal protein S24) as a ribosomal protein, multiple transcript variant encoding different isoforms have been found for this gene. Our previous studies have demonstrated that RPS24 is overexpressed in CRC. However, the mechanisms underlying the role of RPS24 in tumor development have not been fully defined. Methods: Expression of RPS24 isoforms and lncRNA MVIH in CRC tissues and cell lines were quantified by real-time PCR or western blotting assay. Endothelial tube formation assay was performed to determine the effect of RPS24 on tumor angiogenesis. The cell viability of HUVEC was determined by MTT assay, and the migration and invasion ability of HUVEC were detected by transwell assay. PGK1 secretion was tested with a specific ELISA kit. Results: Here, we found that RPS24c isoform was a major contributor to tumor angiogenesis, a vital process in tumor growth and metastasis. Real-time PCR revealed that RPS24c isoform was highly expressed in CRC tissues, while other isoforms are present in both normal and CRC tissues with no statistical difference. Moreover the change of RPS24 protein level is mainly due to the fluctuation of RPS24c. Furthermore, we observed that silencing RPS24c could decrease angiogenesis by inhibiting tubule formation, HUVEC cell proliferation and migration. Additionally, we investigated the molecular mechanisms and demonstrated that RPS24c mRNA interacted with lncRNA MVIH, the binding-interaction enhanced the stability of each other, thereby activated angiogenesis by inhibiting the secretion of PGK1. Conclusion: RPS24c facilitates tumor angiogenesis via the RPS24c/MVIH/PGK1 pathway in CRC. RPS24c inhibition may be a novel option for anti-vascular treatment in CRC.

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Measurement of frequency characteristics of disturbance torque in the process of satellite antenna rotation

International Journal of Applied Electromagnetics and Mechanics ◽

10.3233/jae-209478 ◽

2020 ◽

Vol 64 (1-4) ◽

pp. 1563-1569

Author(s):

Bo Gao ◽

Minglong Xu

Keyword(s):

Real Time ◽

Signal Transmission ◽

Frequency Characteristics ◽

Structural Vibration ◽

Driving Mechanism ◽

Satellite Antenna ◽

Satellite Applications ◽

Study Characteristics ◽

Satellite Signal ◽

Disturbance Torque

As an important equipment for satellite signal reception and transmission, the satellite antenna needs to be rotated in real time to achieve real-time tracking of the target and complete signal transmission during applications. Antenna driving mechanism is generally composed of motor and other components, which will cause some structural vibration during rotation. For high-stability satellite applications, the vibration disturbance torque is a major factor affecting the satellite stability. In order to study characteristics of the disturbance torque, the disturbance data from the antenna under different rotation conditions should be measured. In this paper, the frequency characteristics of disturbance torque of a rotating satellite antenna using stepper motor as driving motor is tested and discussed.

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Groundwater Monitoring Systems to Understand Sea Water Intrusion Dynamics in the Mediterranean: The Neretva Valley and the Southern Venice Coastal Aquifers Case Studies

Water ◽

10.3390/w13040561 ◽

2021 ◽

Vol 13 (4) ◽

pp. 561

Author(s):

Ivan Lovrinović ◽

Alessandro Bergamasco ◽

Veljko Srzić ◽

Chiara Cavallina ◽

Danko Holjević ◽

...

Keyword(s):

Real Time ◽

Coastal Aquifers ◽

Sea Water ◽

Quantitative Information ◽

Water Dynamics ◽

Monitoring Systems ◽

Driving Mechanism ◽

Sea Water Intrusion ◽

Water Intrusion ◽

Aquifer Systems

Sea water intrusion (SWI) has been widely recognized as a global problem, significantly influencing coastal aquifers, mostly through reduced water quality and agricultural production indicators. In this paper, we present the outcomes of the implementation of two independent real-time monitoring systems, planned and installed to get insights on groundwater dynamics within the adjacent coastal aquifer systems, one located in the Neretva Valley, southeastern Croatia, the other located south of the Venice lagoon, northeastern Italy. Both systems are presented with technical details and the capacity to observe, store, and transmit (Neretva site) observed values in real-time. Analysis of time series reveals the significant influence of the sea level oscillations onto the observed groundwater electrical conductivity (EC) and piezometric head values, while precipitation rate is detected as a driving mechanism for groundwater parameters in shallow geological units. The installed monitoring systems are shown to be of great importance to provide qualitative and quantitative information on the processes influencing groundwater and surface water dynamics within two coastal systems.

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Real-time quantitative PCR assay on bacterial DNA: In a model soil system and environmental samples

The Journal of General and Applied Microbiology ◽

10.2323/jgam.49.101 ◽

2003 ◽

Vol 49 (2) ◽

pp. 101-109 ◽

Cited By ~ 15

Author(s):

Shaila Kabir ◽

Narasimmalu Rajendran ◽

Takashi Amemiya ◽

Kiminori Itoh

Keyword(s):

Real Time ◽

Quantitative Pcr ◽

Environmental Samples ◽

Pcr Assay ◽

Soil System ◽

Bacterial Dna ◽

Model Soil ◽

Real Time Quantitative Pcr ◽

Quantitative Pcr Assay

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Carbon Materials Advancing Microorganisms in Driving Soil Organic Carbon Regulation

Research ◽

10.34133/2022/9857374 ◽

2022 ◽

Vol 2022 ◽

pp. 1-12

Author(s):

Chunyu Tang ◽

Fan Yang ◽

Markus Antonietti

Keyword(s):

Soil Carbon ◽

Biological Diversity ◽

Carbon Materials ◽

Carbon Pool ◽

Soil Microbiome ◽

Special Focus ◽

Soil System ◽

Carbon Regulation ◽

Designed Materials ◽

And Control

Carbon emission from soil is not only one of the major sources of greenhouse gases but also threatens biological diversity, agricultural productivity, and food security. Regulation and control of the soil carbon pool are political practices in many countries around the globe. Carbon pool management in engineering sense is much bigger and beyond laws and monitoring, as it has to contain proactive elements to restore active carbon. Biogeochemistry teaches us that soil microorganisms are crucial to manage the carbon content effectively. Adding carbon materials to soil is thereby not directly sequestration, as interaction of appropriately designed materials with the soil microbiome can result in both: metabolization and thereby nonsustainable use of the added carbon, or—more favorably—a biological amplification of human efforts and sequestration of extra CO2 by microbial growth. We review here potential approaches to govern soil carbon, with a special focus set on the emerging practice of adding manufactured carbon materials to control soil carbon and its biological dynamics. Notably, research on so-called “biochar” is already relatively mature, while the role of artificial humic substance (A-HS) in microbial carbon sequestration is still in the developing stage. However, it is shown that the preparation and application of A-HS are large biological levers, as they directly interact with the environment and community building of the biological soil system. We believe that A-HS can play a central role in stabilizing carbon pools in soil.

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Potential pitfalls in the quantitative molecular detection ofEscherichia coliO157:H7 in environmental matrices

Canadian Journal of Microbiology ◽

10.1139/w05-149 ◽

2006 ◽

Vol 52 (5) ◽

pp. 482-488 ◽

Cited By ~ 25

Author(s):

Rebekka R.E Artz ◽

Lisa M Avery ◽

Davey L Jones ◽

Ken Killham

Keyword(s):

Escherichia Coli ◽

Real Time ◽

Real Time Pcr ◽

Environmental Samples ◽

Detection Sensitivity ◽

Escherichia Coli O157 ◽

Soil System ◽

E Coli ◽

Animal Wastes ◽

Pcr Assays

The detection sensitivity and potential interference factors of a commonly used assay based on real-time polymerase chain reaction (PCR) for Escherichia coli O157:H7 using eae gene-specific primers were assessed. Animal wastes and soil samples were spiked with known replicate quantities of a nontoxigenic strain of E. coli O157:H7 in a viable or dead state and as unprotected DNA. The detection sensitivity and accuracy of real-time PCR for E. coli O157:H7 in animal wastes and soil is low compared to enrichment culturing. Nonviable cells and unprotected DNA were shown to produce positive results in several of the environmental samples tested, leading to potential overestimates of cell numbers due to prolonged detection of nonviable cells. This demonstrates the necessity for the specific calibration of real-time PCR assays in environmental samples. The accuracy of the eae gene–based detection method was further evaluated over time in a soil system against an activity measurement, using the bioluminescent properties of an E. coli O157:H7 Tn5luxCDABE construct. The detection of significant numbers of viable but nonculturable (VBNC) as well as nonviable and possibly physically protected cells as shown over a period of 90 days further complicates the use of real-time PCR assays for quick diagnostics in environmental samples and infers that enrichment culturing is still required for the final verification of samples found positive by real-time PCR methods.Key words: Escherichia coli O157:H7, real-time PCR, animal waste, soil, VBNC.

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Real-time dynamics of mutagenesis reveal the chronology of DNA repair and damage tolerance responses in single cells

Proceedings of the National Academy of Sciences ◽

10.1073/pnas.1801101115 ◽

2018 ◽

Vol 115 (28) ◽

pp. E6516-E6525 ◽

Cited By ~ 23

Author(s):

Stephan Uphoff

Keyword(s):

Dna Repair ◽

Real Time ◽

Molecular Mechanisms ◽

Damage Tolerance ◽

Single Cells ◽

Dna Alkylation ◽

Alkylation Damage ◽

Time Dynamics ◽

Dna Mismatches ◽

The Creation

Evolutionary processes are driven by diverse molecular mechanisms that act in the creation and prevention of mutations. It remains unclear how these mechanisms are regulated because limitations of existing mutation assays have precluded measuring how mutation rates vary over time in single cells. Toward this goal, I detected nascent DNA mismatches as a proxy for mutagenesis and simultaneously followed gene expression dynamics in single Escherichia coli cells using microfluidics. This general microscopy-based approach revealed the real-time dynamics of mutagenesis in response to DNA alkylation damage and antibiotic treatments. It also enabled relating the creation of DNA mismatches to the chronology of the underlying molecular processes. By avoiding population averaging, I discovered cell-to-cell variation in mutagenesis that correlated with heterogeneity in the expression of alternative responses to DNA damage. Pulses of mutagenesis are shown to arise from transient DNA repair deficiency. Constitutive expression of DNA repair pathways and induction of damage tolerance by the SOS response compensate for delays in the activation of inducible DNA repair mechanisms, together providing robustness against the toxic and mutagenic effects of DNA alkylation damage.

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Using 13C to Unravel the Molecular Mechanisms of Microbiome Response to Rewetting in Soil

10.5194/egusphere-egu21-13880 ◽

2021 ◽

Author(s):

Mary Lipton ◽

Montana Smith ◽

James Moran ◽

Allison Thompson ◽

Karl Weitz ◽

...

Keyword(s):

Mass Spectrometry ◽

Real Time ◽

Metabolic Pathway ◽

Molecular Mechanisms ◽

Isotope Ratio Mass Spectrometry ◽

Terrestrial Ecosystems ◽

Middle Layer ◽

Nutrient Utilization ◽

Nutrient Cycles ◽

Water Control

Hydration levels influence carbon and nutrient cycles in soils. The rapid carbon release and nutrient utilization upon hydration in desiccated soils has been observed for decades, but little is known about the controls and timing of the underlying molecular events. This research is aimed at developing and using novel isotope based techniques to map the interdependence of carbon and nutrient cycling in soils using 13C labeled substrates as a tracer of nutrient fluxes in desiccated soil microbiomes after rewetting in combination with our novel Real Time Mass Spectrometry (RTMS) system 1, isotope ratio mass spectrometry (IRMS) and omics measurements.Our experiments involve mapping the initial microbial response to wetting, by using a combination of an atmospheric monitoring RTMS instrument that measures the levels of CO2, O2, N2, H2O and their isotopologues, simultaneously in real time and IRMS which will be used to determine the biological fate of the carbon from each substrate. In each experiment desiccated soil that has been kept at drought conditions for 2 weeks was placed in the incubation chamber. Deionized water (control), 13C glucose and 13C alanine dissolved in water was added to the soil through a syringe to mimic a rewetting event. An identical initial hydration response measured by tracing the production of 12CO2 every 5 sec for the first 10 minutes after the addition of water was seen for all the samples potentially indicating that the carbon respired in this initial burst had formed an association with the cell (either intracellular or EPS bound) during dry down. The metabolism and respiration of glucose and alanine, measured by the production of 13CO2 occurred at a much slower time frame (60 to 90 minutes) where the rate of 13CO2 production of the glucose was about 10x that of alanine.IRMS measurements were used to determine the preferential metabolic pathway of each substrate. The soil was removed from the chamber, treated with a mixture of methanol/chloroform/water, beadbeat, sonicated and centrifuged in a modified Folch extraction.&#160; The resulting supernatant was allowed to separate into 3 fractions of polar metabolites (methanol layer), proteins (middle layer) and lipids (chloroform layer).&#160; Each fraction was analyzed by IRMS to quantify the extent of 13C label incorporation into the soil phase to help guide interpretation of the 13CO2 production rates (measured via RTMS), the preferential metabolic pathway of each substrate will be determined. While it was seen that glucose was taken up into all of the biomass partitions as well as respired into CO2, alanine was metabolized to a lesser extent and was predominantly used in protein synthesis.These studies represent approaches that showcase advanced stable isotope analyses to determine molecular provenance in ecosystems by developing an understanding of the molecular mechanisms involved in carbon and nutrient cycles in terrestrial ecosystems.

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Revealing the role of phospholipase Cβ3 in the regulation of VEGF-induced vascular permeability

Blood ◽

10.1182/blood-2012-03-417824 ◽

2012 ◽

Vol 120 (11) ◽

pp. 2167-2173 ◽

Cited By ~ 21

Author(s):

Luke H. Hoeppner ◽

Kathryn N. Phoenix ◽

Karl J. Clark ◽

Resham Bhattacharya ◽

Xun Gong ◽

...

Keyword(s):

Vascular Permeability ◽

Real Time ◽

Molecular Mechanisms ◽

Negative Regulator ◽

Transgenic Zebrafish ◽

Genetic Studies ◽

In Vivo Models ◽

Ischemic Diseases

AbstractVEGF induces vascular permeability (VP) in ischemic diseases and cancer, leading to many pathophysiological consequences. The molecular mechanisms by which VEGF acts to induce hyperpermeability are poorly understood and in vivo models that easily facilitate real-time, genetic studies of VP do not exist. In the present study, we report a heat-inducible VEGF transgenic zebrafish (Danio rerio) model through which VP can be monitored in real time. Using this approach with morpholino-mediated gene knock-down and knockout mice, we describe a novel role of phospholipase Cβ3 as a negative regulator of VEGF-mediated VP by regulating intracellular Ca2+ release. Our results suggest an important effect of PLCβ3 on VP and provide a new model with which to identify genetic regulators of VP crucial to several disease processes.

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A Promising Antiprion Trimethoxychalcone Binds to the Globular Domain of the Cellular Prion Protein and Changes Its Cellular Location

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.01441-17 ◽

2017 ◽

Vol 62 (2) ◽

Cited By ~ 6

Author(s):

N. C. Ferreira ◽

L. M. Ascari ◽

A. G. Hughson ◽

G. R. Cavalheiro ◽

C. F. Góes ◽

...

Keyword(s):

Cell Surface ◽

Real Time ◽

Prion Protein ◽

Molecular Mechanisms ◽

Cellular Prion Protein ◽

Transmissible Spongiform Encephalopathies ◽

Globular Domain ◽

Mrna Levels ◽

Spongiform Encephalopathies

ABSTRACTThe search for antiprion compounds has been encouraged by the fact that transmissible spongiform encephalopathies (TSEs) share molecular mechanisms with more prevalent neurodegenerative pathologies, such as Parkinson's and Alzheimer's diseases. Cellular prion protein (PrPC) conversion into protease-resistant forms (protease-resistant PrP [PrPRes] or the scrapie form of PrP [PrPSc]) is a critical step in the development of TSEs and is thus one of the main targets in the screening for antiprion compounds. In this work, three trimethoxychalcones (compounds J1, J8, and J20) and one oxadiazole (compound Y17), previously identifiedin vitroto be potential antiprion compounds, were evaluated through different approaches in order to gain inferences about their mechanisms of action. None of them changed PrPCmRNA levels in N2a cells, as shown by reverse transcription-quantitative real-time PCR. Among them, J8 and Y17 were effective in real-time quaking-induced conversion reactions using rodent recombinant PrP (rPrP) from residues 23 to 231 (rPrP23–231) as the substrate and PrPScseeds from hamster and human brain. However, when rPrP from residues 90 to 231 (rPrP90–231), which lacks the N-terminal domain, was used as the substrate, only J8 remained effective, indicating that this region is important for Y17 activity, while J8 seems to interact with the PrPCglobular domain. J8 also reduced the fibrillation of mouse rPrP23–231seeded within vitro-produced fibrils. Furthermore, most of the compounds decreased the amount of PrPCon the N2a cell surface by trapping this protein in the endoplasmic reticulum. On the basis of these results, we hypothesize that J8, a nontoxic compound previously shown to be a promising antiprion agent, may act by different mechanisms, since its efficacy is attributable not only to PrP conversion inhibition but also to a reduction of the PrPCcontent on the cell surface.

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