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1. The Link between Purine Metabolism and Production of Antibiotics in Streptomyces

   https://doi.org/10.3390/antibiotics8020076  

   Sivapragasam, Smitha; Grove, Anne (June 2019, Antibiotics)

   Stress and starvation causes bacterial cells to activate the stringent response. This results in down-regulation of energy-requiring processes related to growth, as well as an upregulation of genes associated with survival and stress responses. Guanosine tetra- and pentaphosphates (collectively referred to as (p)ppGpp) are critical for this process. In Gram-positive bacteria, a main function of (p)ppGpp is to limit cellular levels of GTP, one consequence of which is reduced transcription of genes that require GTP as the initiating nucleotide, such as rRNA genes. In Streptomycetes, the stringent response is also linked to complex morphological differentiation and to production of secondary metabolites, including antibiotics. These processes are also influenced by the second messenger c-di-GMP. Since GTP is a substrate for both (p)ppGpp and c-di-GMP, a finely tuned regulation of cellular GTP levels is required to ensure adequate synthesis of these guanosine derivatives. Here, we discuss mechanisms that operate to control guanosine metabolism and how they impinge on the production of antibiotics in Streptomyces species. 

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2. Cell cycle control and environmental response by second messengers in Caulobacter crescentus

   https://doi.org/10.1186/s12859-020-03687-z  

   Xu, Chunrui; Weston, Bronson R.; Tyson, John J.; Cao, Yang (September 2020, BMC Bioinformatics)

   null (Ed.)

   Abstract Background Second messengers, c-di-GMP and (p)ppGpp, are vital regulatory molecules in bacteria, influencing cellular processes such as biofilm formation, transcription, virulence, quorum sensing, and proliferation. While c-di-GMP and (p)ppGpp are both synthesized from GTP molecules, they play antagonistic roles in regulating the cell cycle. In C. crescentus , c-di-GMP works as a major regulator of pole morphogenesis and cell development. It inhibits cell motility and promotes S-phase entry by inhibiting the activity of the master regulator, CtrA. Intracellular (p)ppGpp accumulates under starvation, which helps bacteria to survive under stressful conditions through regulating nucleotide levels and halting proliferation. (p)ppGpp responds to nitrogen levels through RelA-SpoT homolog enzymes, detecting glutamine concentration using a nitrogen phosphotransferase system (PTS Ntr ). This work relates the guanine nucleotide-based second messenger regulatory network with the bacterial PTS Ntr system and investigates how bacteria respond to nutrient availability. Results We propose a mathematical model for the dynamics of c-di-GMP and (p)ppGpp in C. crescentus and analyze how the guanine nucleotide-based second messenger system responds to certain environmental changes communicated through the PTS Ntr system. Our mathematical model consists of seven ODEs describing the dynamics of nucleotides and PTS Ntr enzymes. Our simulations are consistent with experimental observations and suggest, among other predictions, that SpoT can effectively decrease c-di-GMP levels in response to nitrogen starvation just as well as it increases (p)ppGpp levels. Thus, the activity of SpoT (or its homologues in other bacterial species) can likely influence the cell cycle by influencing both c-di-GMP and (p)ppGpp. Conclusions In this work, we integrate current knowledge and experimental observations from the literature to formulate a novel mathematical model. We analyze the model and demonstrate how the PTS Ntr system influences (p)ppGpp, c-di-GMP, GMP and GTP concentrations. While this model does not consider all aspects of PTS Ntr signaling, such as cross-talk with the carbon PTS system, here we present our first effort to develop a model of nutrient signaling in C. crescentus . 

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3. The nucleotide pGpp acts as a third alarmone in Bacillus, with functions distinct from those of (p)ppGpp

   https://doi.org/10.1038/s41467-020-19166-1  

   Yang, Jin; Anderson, Brent W.; Turdiev, Asan; Turdiev, Husan; Stevenson, David M.; Amador-Noguez, Daniel; Lee, Vincent T.; Wang, Jue D. (December 2020, Nature Communications)

   null (Ed.)

   Abstract The alarmone nucleotides guanosine tetraphosphate and pentaphosphate, commonly referred to as (p)ppGpp, regulate bacterial responses to nutritional and other stresses. There is evidence for potential existence of a third alarmone, guanosine-5′-monophosphate-3′-diphosphate (pGpp), with less-clear functions. Here, we demonstrate the presence of pGpp in bacterial cells, and perform a comprehensive screening to identify proteins that interact respectively with pGpp, ppGpp and pppGpp in Bacillus species. Both ppGpp and pppGpp interact with proteins involved in inhibition of purine nucleotide biosynthesis and with GTPases that control ribosome assembly or activity. By contrast, pGpp interacts with purine biosynthesis proteins but not with the GTPases. In addition, we show that hydrolase NahA (also known as YvcI) efficiently produces pGpp by hydrolyzing (p)ppGpp, thus modulating alarmone composition and function. Deletion of nahA leads to reduction of pGpp levels, increased (p)ppGpp levels, slower growth recovery from nutrient downshift, and loss of competitive fitness. Our results support the existence and physiological relevance of pGpp as a third alarmone, with functions that can be distinct from those of (p)ppGpp. 

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4. Temporal regulation of cell polarity via the interaction of the Ras GTPase Rsr1 and the scaffold protein Bem1

   https://doi.org/10.1091/mbc.E19-02-0106  

   Miller, Kristi E.; Lo, Wing-Cheong; Chou, Ching-Shan; Park, Hay-Oak; Li, Rong (September 2019, Molecular Biology of the Cell)

   The Cdc42 guanosine triphosphatase (GTPase) plays a central role in polarity development in species ranging from yeast to humans. In budding yeast, a specific growth site is selected in the G1 phase. Rsr1, a Ras GTPase, interacts with Cdc42 and its associated proteins to promote polarized growth at the proper bud site. Yet how Rsr1 regulates cell polarization is not fully understood. Here, we show that Rsr1-GDP interacts with the scaffold protein Bem1 in early G1, likely hindering the role of Bem1 in Cdc42 polarization and polarized secretion. Consistent with these in vivo observations, mathematical modeling predicts that Bem1 is unable to promote Cdc42 polarization in early G1 in the presence of Rsr1-GDP. We find that a part of the Bem1 Phox homology domain, which overlaps with a region interacting with the exocyst component Exo70, is necessary for the association of Bem1 with Rsr1-GDP. Overexpression of the GDP-locked Rsr1 interferes with Bem1-dependent Exo70 polarization. We thus propose that Rsr1 functions in spatial and temporal regulation of polarity establishment by associating with distinct polarity factors in its GTP- and GDP-bound states. 

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5. Formation and Measurement of Ras Y32 Mutant's Role in GTP Hydrolysis

   Heard, J.; Landry, D.; Novelli, E.; Webb, L. (August 2019, 2019 BMES Conference Proceedings - REU Abstract Accepted Poster)

   Introduction: Members of the Ras protein family are involved in sending signals through the human body to regulate processes including cell growth. Acting as a molecular switch, Ras alternates between its active and inactive states (“on” and “off”) depending on its binding to either GTP or GDP. Upon the introduction of a mutation, Ras becomes incapable of performing the GTP hydrolysis reaction at a controlled rate. This results in the protein remaining in the “on” state for prolonged periods of time, disturbing its role in the regulation of cell growth and causing tumors to form. In the genome of cancerous tumors, mutations that permanently activate Ras have been observed and analyzed at three distinct locations, one being Q61. Previous research1 shows how the Q61 residue contributes to the protein’s function by stabilizing the water molecule in the active site, but little research has been done studying the Y32 residue, which has also been implicated in the mechanism. This research investigates the role Y32 plays in activating Ras via introduction of a different amino acid and measurement of the mutated protein’s activity in the hydrolysis of GTP. 

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