scholarly journals Substrate dependent homeostatic control of c-di-AMP synthase (MsDisA) and hydrolase (MsPDE) from Mycobacterium smegmatis

2021 ◽  
Author(s):  
Sudhanshu Gautam ◽  
Avisek Mahapa ◽  
Lahari Yeramala ◽  
Apoorv Gandhi ◽  
Sushma Krishnan ◽  
...  
Keyword(s):  
Stress Response ◽  
Mycobacterium Smegmatis ◽  
Thermotoga Maritima ◽  
Structural Characteristics ◽  
Second Messengers ◽  
Alkaline Stress ◽  
The Past ◽  
Physiological Processes ◽  
Biochemical Assays ◽  
Minimal Region

In bacteria, cyclic-di-nucleotide based second messengers regulate various physiological processes including the stress response. For the past few decades, cyclic diadenosine monophosphate (c-di-AMP) is emerging as a crucial second messenger in bacterial world. It being an essential molecule, is implicated in fatty acid metabolism, antibiotic resistance, biofilm formation, virulence and activates the cytosolic pathway of innate immunity in host cell. The level of c-di-AMP is maintained within the cell by the action of two opposing enzymes, namely diadenylate cyclases and phosphodiesterases. However, such kind of c-di-AMP modulation remains to be explored in Mycobacterium smegmatis. Here, we systematically investigate the c-di-AMP synthase (MsDisA) and a hydrolase (MsPDE) from M. smegmatis at different pHs and osmolytic conditions. Our biochemical assays showed that the MsDisA activity is enhanced during the alkaline stress and c-di-AMP is readily produced without any intermediates. At pH 9.4, the MsDisA promoter activity increases significantly, further strengthening this observation. However, under physiological conditions, the activity of MsDisA is moderate with the formation of intermediates. We also observe that the size of MsDisA is significantly increased upon incubation with substrate. To further get deep insights into the structural characteristics, we report a 3.8 Å cryo-EM structure of the MsDisA protein, distinct from the earlier reported structure of DisA from Thermotoga maritima. The domain mutant experiments prove that the N-terminal minimal region can form a functional octamer. Thus, our results reveal how mycobacterial c-di-AMP is biochemically and structurally regulated in response to different environments. Keywords: Mycobacteria, second messengers, stress response, Cyclic-di-AMP, MsDisA, MsPDE, Cryo-EM

scholarly journals RelZ-Mediated Stress Response in Mycobacterium smegmatis: pGpp Synthesis and Its Regulation

2019 ◽  
Vol 202 (2) ◽  
Author(s):  
Anushya Petchiappan ◽  
Sujay Y. Naik ◽  
Dipankar Chatterji
Keyword(s):  
Stress Response ◽  
Mycobacterium Smegmatis ◽  
Biofilm Formation ◽  
Second Messengers ◽  
Antibiotic Tolerance ◽  
Response Mechanism ◽  
Term Survival ◽  
Content Type ◽  
Guanosine Tetraphosphate

ABSTRACT Stringent response is a conserved stress response mechanism in which bacteria employ the second messengers guanosine tetraphosphate and guanosine pentaphosphate [collectively termed (p)ppGpp] to reprogram their cellular processes under stress. In mycobacteria, these alarmones govern a multitude of cellular phenotypes, such as cell division, biofilm formation, antibiotic tolerance, and long-term survival. Mycobacterium smegmatis possesses the bifunctional RelMsm as a (p)ppGpp synthetase and hydrolase. In addition, it contains a short alarmone synthetase MS_RHII-RSD (renamed RelZ), which contains an RNase H domain in tandem with the (p)ppGpp synthetase domain. The physiological functions of RelMsm have been well documented, but there is no clear picture about the cellular functions of RelZ in M. smegmatis. RelZ has been implicated in R-loop induced stress response due to its unique domain architecture. In this study, we elucidate the differential substrate utilization pattern of RelZ compared to that of RelMsm. We unveil the ability of RelZ to use GMP as a substrate to synthesize pGpp, thereby expanding the repertoire of second messengers known in mycobacteria. We have demonstrated that the pGpp synthesis activity of RelZ is negatively regulated by RNA and pppGpp. Furthermore, we investigated its role in biofilm formation and antibiotic tolerance. Our findings highlight the complex role played by the RelZ in cellular physiology of M. smegmatis and sheds light upon its functions distinct from those of RelMsm. IMPORTANCE Bacteria utilize nucleotide messengers to survive the hostile environmental conditions and the onslaught of attacks within the host. The second messengers guanosine tetraphosphate and pentaphosphate [(p)ppGpp] have a profound impact on the long-term survival, biofilm formation, antibiotic tolerance, virulence, and pathogenesis of bacteria. Therefore, understanding the stress response mechanism regulated by (p)ppGpp is essential for discovering inhibitors of stress response and potential drug targets. Mycobacterium smegmatis contains two (p)ppGpp synthetases: RelMsm and RelZ. Our study unravels the novel regulatory mechanisms of RelZ activity and its role in mediating antibiotic tolerance. We further reveal its ability to synthesize novel second messenger pGpp, which may have regulatory roles in mycobacteria.

scholarly journals Alternative approaches to target Myc for cancer treatment

2021 ◽  
Vol 6 (1) ◽  
Author(s):  
Chen Wang ◽  
Jiawei Zhang ◽  
Jie Yin ◽  
Yichao Gan ◽  
Senlin Xu ◽  
...  
Keyword(s):  
Cancer Treatment ◽  
Small Molecules ◽  
Drug Target ◽  
The Past ◽  
Physiological Processes ◽  
Alternative Approaches ◽  
Global Transcriptome ◽  
Factor C ◽  
Signaling Modules ◽  
Druggable Targets

AbstractThe Myc proto-oncogene family consists of three members, C-MYC, MYCN, and MYCL, which encodes the transcription factor c-Myc (hereafter Myc), N-Myc, and L-Myc, respectively. Myc protein orchestrates diverse physiological processes, including cell proliferation, differentiation, survival, and apoptosis. Myc modulates about 15% of the global transcriptome, and its deregulation rewires the cellular signaling modules inside tumor cells, thereby acquiring selective advantages. The deregulation of Myc occurs in >70% of human cancers, and is related to poor prognosis; hence, hyperactivated Myc oncoprotein has been proposed as an ideal drug target for decades. Nevertheless, no specific drug is currently available to directly target Myc, mainly because of its “undruggable” properties: lack of enzymatic pocket for conventional small molecules to bind; inaccessibility for antibody due to the predominant nucleus localization of Myc. Although the topic of targeting Myc has actively been reviewed in the past decades, exciting new progresses in this field keep emerging. In this review, after a comprehensive summarization of valuable sources for potential druggable targets of Myc-driven cancer, we also peer into the promising future of utilizing macropinocytosis to deliver peptides like Omomyc or antibody agents to intracellular compartment for cancer treatment.

scholarly journals Tree Vitality and Forest Health: Can Tree-Ring Stable Isotopes Be Used as Indicators?

2021 ◽  
Author(s):  
Paolo Cherubini ◽  
Giovanna Battipaglia ◽  
John L. Innes
Keyword(s):  
Air Pollution ◽  
Stable Isotopes ◽  
Tree Ring ◽  
Forest Decline ◽  
Forest Health ◽  
Stress Factors ◽  
Tree Vitality ◽  
The Past ◽  
Physiological Processes

Abstract Purpose of Review Society is concerned about the long-term condition of the forests. Although a clear definition of forest health is still missing, to evaluate forest health, monitoring efforts in the past 40 years have concentrated on the assessment of tree vitality, trying to estimate tree photosynthesis rates and productivity. Used in monitoring forest decline in Central Europe since the 1980s, crown foliage transparency has been commonly believed to be the best indicator of tree condition in relation to air pollution, although annual variations appear more closely related to water stress. Although crown transparency is not a good indicator of tree photosynthesis rates, defoliation is still one of the most used indicators of tree vitality. Tree rings have been often used as indicators of past productivity. However, long-term tree growth trends are difficult to interpret because of sampling bias, and ring width patterns do not provide any information about tree physiological processes. Recent Findings In the past two decades, tree-ring stable isotopes have been used not only to reconstruct the impact of past climatic events, such as drought, but also in the study of forest decline induced by air pollution episodes, and other natural disturbances and environmental stress, such as pest outbreaks and wildfires. They have proven to be useful tools for understanding physiological processes and tree response to such stress factors. Summary Tree-ring stable isotopes integrate crown transpiration rates and photosynthesis rates and may enhance our understanding of tree vitality. They are promising indicators of tree vitality. We call for the use of tree-ring stable isotopes in future monitoring programmes.

scholarly journals VTC4 Polyphosphate Polymerase Knockout Increases Stress Resistance of Saccharomyces cerevisiae Cells

Biology ◽  
2021 ◽  
Vol 10 (6) ◽  
pp. 487
Author(s):  
Alexander Tomashevsky ◽  
Ekaterina Kulakovskaya ◽  
Ludmila Trilisenko ◽  
Ivan V. Kulakovskiy ◽  
Tatiana Kulakovskaya ◽  
...  
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Heavy Metal ◽  
Stress Response ◽  
Stress Resistance ◽  
Alkaline Stress ◽  
Inorganic Polyphosphate ◽  
Divalent Metal Transporter ◽  
Metal Transporter ◽  
Elevated Expression ◽  
Chaperone Complex

Inorganic polyphosphate (polyP) is an important factor of alkaline, heavy metal, and oxidative stress resistance in microbial cells. In yeast, polyP is synthesized by Vtc4, a subunit of the vacuole transporter chaperone complex. Here, we report reduced but reliably detectable amounts of acid-soluble and acid-insoluble polyPs in the Δvtc4 strain of Saccharomyces cerevisiae, reaching 10% and 20% of the respective levels of the wild-type strain. The Δvtc4 strain has decreased resistance to alkaline stress but, unexpectedly, increased resistance to oxidation and heavy metal excess. We suggest that increased resistance is achieved through elevated expression of DDR2, which is implicated in stress response, and reduced expression of PHO84 encoding a phosphate and divalent metal transporter. The decreased Mg2+-dependent phosphate accumulation in Δvtc4 cells is consistent with reduced expression of PHO84. We discuss a possible role that polyP level plays in cellular signaling of stress response mobilization in yeast.

Investigation by NMR spectroscopy of the structural characteristics of modified oligo-nucleotides with pendant porphyrins

2019 ◽  
Vol 23 (07n08) ◽  
pp. 797-812 ◽  
Author(s):  
Sonja Merkaš ◽  
Mladen Žinić ◽  
Régis Rein ◽  
Nathalie Solladié
Keyword(s):  
Nmr Spectroscopy ◽  
Structural Characteristics ◽  
Strong Interactions ◽  
Light Harvesting Complexes ◽  
The Past ◽  
Naturally Occurring ◽  
Nmr Data ◽  
Intermolecular Distances ◽  
Uracil Base ◽  
Biopolymer Scaffolds

During the past years, we focused on exerting control over the position and distance of porphyrins along our specifically designed oligonucleotidic scaffold. Indeed, in naturally occurring light-harvesting complexes, biopolymer scaffolds hold pigments at intermolecular distances that optimize photon capture, electronic coupling, and energy transfer. To this end, four uridine-porphyrin conjugates (a monomer, a dimer, a tetramer and an octamer) were subjected to a comprehensive conformational analysis by using NMR spectroscopy. The collected NOE NMR data highlighted characteristic and strong interactions indicating that the glycosidic angle between the ribose and uracil base is anti. In order to further investigate the conformation of this family of molecules, NMR experiments were carried out at variable temperatures. At low temperature, the signals of the porphyrinic protons decoalesce, showing two sets of [Formula: see text]-pyrrolic protons. Similar observations are made for signals corresponding to sugar moieties and especially the H1′ protons, indicating molecular motions within our porphyrin-uridin arrays. These results testify in favor of the existence of a dynamic process between C3′-endo and C2′-endo conformations.

The structure and changes of China’s land system

2019 ◽  
Vol 11 (3) ◽  
pp. 471-488 ◽  
Author(s):  
Liu Shouying
Keyword(s):  
Economic Growth ◽  
Institutional Change ◽  
Structural Changes ◽  
Structural Characteristics ◽  
Institutional Arrangements ◽  
Reform Process ◽  
Content Type ◽  
The Past ◽  
Land System ◽  
Rapid Economic Growth

Purpose The purpose of this paper is to analyze the structure and changes of China’s land system. To achieve this aim, the paper is divided into four parts. The first part gives a brief introduction to the structural characteristics of the Chinese land institutional arrangements; the second part analyzes the reform process of the land system in the past 40 years and its path of change; the third part engages the discussion about the historic contribution made by the land institutional change to rapid economic growth and structural changes; and the final part is conclusion and some policy implications. Design/methodology/approach After 40 years of reforms and opening up, China has not only created a growth miracle unparalleled for any major country in human history, but also transformed itself from a rural to an urban society. Behind this great transformation is a systemic reform in land institutions. Rural land institutions went from collectively owned to household responsibility system, thereby protecting farmers’ land rights. This process resulted in long-term sustainable growth in China’s agriculture, a massive rural-urban migration and a historical agricultural transformation. The conversion of agricultural land to non-agricultural uses and the introduction of market mechanisms made land a policy tool in driving high economic growth, industrialization and urbanization. Findings Research shows that the role of land and its relationship with the economy will inevitably change as China’s economy enters a new stage of medium-to-high speed growth. With economic restructuring, low-cost industrial land will be less effective. Urbanization is also shifting from rapid expansion to endogenous growth so that returns on land capitalization will decrease and risks will increase. Therefore, China must abandon land-dependent growth model through deepening land reforms and adapt a new pattern of economic development. Originality/value This paper gives a brief introduction to the structural characteristics of the Chinese land institutional arrangements, analyzes the reform process of the land system in the past 40 years and its path of change, and evaluates the historic contribution made by the land institutional change to rapid economic growth and structural changes.

scholarly journals Phosphodiesterase Activities in the Eye of Old and Young Rats in Normoxic, Hypoxic and Hyperoxic Atmospheres

2003 ◽  
Vol 1 (1) ◽  
pp. 25-32 ◽  
Author(s):  
G. Spoto ◽  
A. Contento ◽  
M. Di Nicola ◽  
G. Bianchi ◽  
C. Di Giulio ◽  
...  
Keyword(s):  
Cyclic Nucleotides ◽  
Biological Effects ◽  
Atmospheric Oxygen ◽  
Second Messengers ◽  
Camp Phosphodiesterase ◽  
Young Rats ◽  
Physiological Processes ◽  
Adaptive Processes ◽  
Old Rats

Phosphodiesterase activity was tested on homogenized eyes of young and old rats kept in hypoxic and hyperoxic conditions, with the aim of correlating any difference in PDE activity with aging and variations in atmospheric oxygen contents. The activities of the two enzymes, cAMP phosphodiesterase (cAMP-PDE) and cGMP phosphodiesterase (cGMP-PDE), were tested. Phosphodiesterases seem to be particularly susceptible to variations in oxygen tension, suggesting an important role of cyclic nucleotides in cellular adaptive processes. Particularly, cAMP-PDE activity increases lightly both in hypoxic and hyperoxic conditions in young and old rats. For cGMP-PDE activity of young rats, a similar behaviour to cAMP-PDE activity is observed with a similar increase in hypoxic and hyperoxic conditions respect to the control rats. Instead old rats seem to be quite insensible to hypoxia, while they show a fair increase in cGMP-PDE activity in the case of hyperoxia. The second messengers cAMP and cGMP play important roles in mediating the biological effects of a wide variety of first messengers. The intracellular levels of cyclic nucleotides depend upon rates of synthesis and degradation, actuated, respectively, by cyclases and phosphodiesterases (PDEs). Therefore, PDEs seem to play an important role in a wide variety of physiological processes.

scholarly journals Make and break the alarmone: regulation of (p)ppGpp synthetase/hydrolase enzymes in bacteria

2019 ◽  
Vol 43 (4) ◽  
pp. 389-400 ◽  
Author(s):  
Séverin Ronneau ◽  
Régis Hallez
Keyword(s):  
Cell Cycle ◽  
Stress Response ◽  
Stress Responses ◽  
Biofilm Formation ◽  
Pathogenic Bacteria ◽  
Second Messengers ◽  
Molecular Devices ◽  
Environmental Cues ◽  
Fluctuating Environments ◽  
Harsh Conditions

ABSTRACTBacteria use dedicated mechanisms to respond adequately to fluctuating environments and to optimize their chances of survival in harsh conditions. One of the major stress responses used by virtually all bacteria relies on the sharp accumulation of an alarmone, the guanosine penta- or tetra-phosphate commonly referred to as (p)ppGpp. Under stressful conditions, essentially nutrient starvation, these second messengers completely reshape the metabolism and physiology by coordinately modulating growth, transcription, translation and cell cycle. As a central regulator of bacterial stress response, the alarmone is also involved in biofilm formation, virulence, antibiotics tolerance and resistance in many pathogenic bacteria. Intracellular concentrations of (p)ppGpp are determined by a highly conserved and widely distributed family of proteins called RelA-SpoT Homologs (RSH). Recently, several studies uncovering mechanisms that regulate RSH activities have renewed a strong interest in this field. In this review, we outline the diversity of the RSH protein family as well as the molecular devices used by bacteria to integrate and transform environmental cues into intracellular (p)ppGpp levels.

Piramal Diagnostics

2011 ◽  
Vol 15 (01) ◽  
pp. 97-121
Author(s):  
Neeraj Dwivedi ◽  
Arvinder Singh
Keyword(s):  
Strategic Planning ◽  
Structural Characteristics ◽  
Vice President ◽  
Medical Diagnostics ◽  
Growth Strategy ◽  
Decision Situation ◽  
The Past ◽  
Strategic Option ◽  
The Future ◽  
Future Growth

The case presents a decision situation facing the Vice President of strategic planning at Piramal Diagnostics Limited, who has to formulate the future growth strategy and decide on the roadmap. The company is the largest player in the organized medical diagnostics industry in India and has shown attractive growth in the past few years. The case describes the structural characteristics of the medical diagnostics industry in India and follows it with a description of the strengths and weaknesses of Piramal Diagnostics and the strategies adopted by it. The Vice President is expected to choose an appropriate strategic option to help the company achieve its ambitious growth target.

scholarly journals An uracil-linked hydroxyflavone probe for the recognition of ATP

2018 ◽  
Vol 14 ◽  
pp. 747-755 ◽  
Author(s):  
Márton Bojtár ◽  
Péter Zoltán Janzsó-Berend ◽  
Dávid Mester ◽  
Dóra Hessz ◽  
Mihály Kállay ◽  
...  
Keyword(s):  
Fluorescence Spectroscopy ◽  
Electrostatic Interactions ◽  
Theoretical Calculations ◽  
Structural Similarity ◽  
Living Systems ◽  
Selective Detection ◽  
Complex Structures ◽  
Fluorescent Chemosensors ◽  
The Past ◽  
Physiological Processes

Background: Nucleotides are essential molecules in living systems due to their paramount importance in various physiological processes. In the past years, numerous attempts were made to selectively recognize and detect these analytes, especially ATP using small-molecule fluorescent chemosensors. Despite the various solutions, the selective detection of ATP is still challenging due to the structural similarity of various nucleotides. In this paper, we report the conjugation of a uracil nucleobase to the known 4’-dimethylamino-hydroxyflavone fluorophore. Results: The complexation of this scaffold with ATP is already known. The complex is held together by stacking and electrostatic interactions. To achieve multi-point recognition, we designed the uracil-appended version of this probe to include complementary base-pairing interactions. The theoretical calculations revealed the availability of multiple complex structures. The synthesis was performed using click chemistry and the nucleotide recognition properties of the probe were evaluated using fluorescence spectroscopy. Conclusions: The first, uracil-containing fluorescent ATP probe based on a hydroxyflavone fluorophore was synthesized and evaluated. A selective complexation with ATP was observed and a ratiometric response in the excitation spectrum.

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