Reverting p53 activation after recovery of cellular stress to resume with cell cycle progression

Publication date: Available online 9 February 2017
Source:Cellular Signalling
Author(s): Pedro A. Lazo
The activation of p53 in response to different types of cellular stress induces several protective reactions including cell cycle arrest, senescence or cell death. These protective effects are a consequence of the activation of p53 by specific phosphorylation performed by several kinases. The reversion of the cell cycle arrest, induced by p53, is a consequence of the phosphorylated and activated p53, which triggers its own downregulation and that of its positive regulators. The different down-regulatory processes have a sequential and temporal order of events. The mechanisms implicated in p53 down-regulation include phosphatases, deacetylases, and protein degradation by the proteasome or autophagy, which also affect different p53 protein targets and functions. The necessary first step is the dephosphorylation of p53 to make it available for interaction with mdm2 ubiquitin-ligase, which requires the activation of phosphatases targeting both p53 and p53-activating kinases. In addition, deacetylation of p53 is required to make lysine residues accessible to ubiquitin ligases. The combined action of these downregulatory mechanisms brings p53 protein back to its basal levels, and cell cycle progression can resume if cells have overcome the stress or damage situation. The specific targeting of these down-regulatory mechanisms can be exploited for therapeutic purposes in cancers harbouring wild-type p53.



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Transcriptional and post-transcriptional regulation of Cdc20 during the spindle assembly checkpoint in S. cerevisiae

Publication date: Available online 9 February 2017
Source:Cellular Signalling
Author(s): Ruiwen Wang, Janet L. Burton, Mark J. Solomon
The anaphase-promoting complex (APC) is a ubiquitin ligase responsible for promoting the degradation of many cell cycle regulators. One of the activators and substrate-binding proteins for the APC is Cdc20. It has been shown previously that Cdc20 can promote its own degradation by the APC in normal cycling cells mainly through a cis-degradation mode (i.e. via an intramolecular mechanism). However, how Cdc20 is degraded during the spindle assembly checkpoint (SAC) is still not fully clear. In this study, we used a dual-Cdc20 system to investigate this issue and found that the cis-degradation mode is also the major pathway responsible for Cdc20 degradation during the SAC. In addition, we found that there is an inverse relationship between APC^Cdc20 activity and the transcriptional activity of the CDC20 promoter, which likely occurs through feedback regulation by APC^Cdc20 substrates, such as the cyclins Clb2 and Clb5. These findings contribute to our understanding of how the inhibition of APC^Cdc20 activity and enhanced Cdc20 degradation are required for proper spindle checkpoint arrest.



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Regulation of inflammation by β-arrestins: Not just receptor Tales

Publication date: Available online 9 February 2017
Source:Cellular Signalling
Author(s): Neil J. Freedman, Sudha K. Shenoy
The ubiquitously expressed, multifunctional scaffolding proteins β-arrestin1 and β-arrestin2 each affect inflammatory signaling in a variety of cell lines. In addition to binding the carboxyl-terminal tails of innumerable 7-transmembrane receptors, β-arrestins scaffold untold numbers of other plasma membrane and cytoplasmic proteins. Consequently, the effects of β-arrestins on inflammatory signaling are diverse, and context-specific. This review highlights the roles of β-arrestins in regulating canonical activation of the pro-inflammatory transcription factor NFκB.



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Transcriptional regulation of RACK1 and modulation of its expression: Role of steroid hormones and significance in health and aging

Publication date: Available online 9 February 2017
Source:Cellular Signalling
Author(s): Erica Buoso, Marilisa Galasso, Melania Maria Serafini, Melania Ronfani, Cristina Lanni, Emanuela Corsini, Marco Racchi
The Receptor for Activated C Kinase 1 (RACK1) is a scaffold protein for different kinases and membrane receptors. RACK1 can shuttle proteins to their sites of action, facilitate cross-talk among distinct signaling pathways or recruit other signaling proteins into the complexes. Therefore, it is a key mediator of various pathways and is involved in various biological events including development, immune response, brain activity and cancer. Because of its importance, it is of extreme significance to understand the transcriptional mechanisms governing its expression. The identification of regulatory elements in the promoter of RACK1 shed some light on its transcriptional modulation in physiological and pathological context. Literature data support the existence of a complex hormonal balance, between glucocorticoids and androgens, in the control of RACK1 expression due to specific and complex interactions on the RACK1 promoter. These and other informations suggest that a better understanding of RACK1 transcriptional regulation is essential to unravel its role. Furthermore, the modulation of its expression in physiological or pathological conditions may be of interest in different context, such as aging and cancer.



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Synthesis of L-cysteine derivatives containing stable sulfur isotopes and application of this synthesis to reactive sulfur metabolome

Publication date: Available online 9 February 2017
Source:Free Radical Biology and Medicine
Author(s): Katsuhiko Ono, Minkyung Jung, Tianli Zhang, Hiroyasu Tsutsuki, Hiroshi Sezaki, Hideshi Ihara, Fan-Yan Wei, Kazuhito Tomizawa, Takaaki Akaike, Tomohiro Sawa
Cysteine persulfide is an L-cysteine derivative having one additional sulfur atom bound to a cysteinyl thiol group, and it serves as a reactive sulfur species that regulates redox homeostasis in cells. Here, we describe a rapid and efficient method of synthesis of L-cysteine derivatives containing isotopic sulfur atoms and application of this method to a reactive sulfur metabolome. We used bacterial cysteine syntheses to incorporate isotopic sulfur atoms into the sulfhydryl moiety of L-cysteine. We cloned three cysteine synthases—CysE, CysK, and CysM—from the Gram-negative bacterium Salmonella enterica serovar Typhimurium LT2, and we generated their recombinant enzymes. We synthesized ³⁴S-labeled L-cysteine from O-acetyl-L-serine and ³⁴S-labeled sodium sulfide as substrates for the CysK or CysM reactions. Isotopic labeling of L-cysteine at both sulfur (³⁴S) and nitrogen (¹⁵N) atoms was also achieved by performing enzyme reactions with ¹⁵N-labeled L-serine, acetyl-CoA, and ³⁴S-labeled sodium sulfide in the presence of CysE and CysK. The present enzyme systems can be applied to syntheses of a series of L-cysteine derivatives including L-cystine, L-cystine persulfide, S-sulfo-L-cysteine, L-cysteine sulfonate, and L-selenocystine. We also prepared ³⁴S-labeled N-acetyl-L-cysteine (NAC) by incubating ³⁴S-labeled L-cysteine with acetyl coenzyme A in test tubes. Tandem mass spectrometric identification of low-molecular-weight thiols after monobromobimane derivatization revealed the endogenous occurrence of NAC in the cultured mammalian cells such as HeLa cells and J774.1 cells. Furthermore, we successfully demonstrated, by using ³⁴S-labeled NAC, metabolic conversion of NAC to glutathione and its persulfide, via intermediate formation of L-cysteine, in the cells. The approach using isotopic sulfur labeling combined with mass spectrometry may thus contribute to greater understanding of reactive sulfur metabolome and redox biology.

Graphical abstract

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How widespread is stable protein S-nitrosylation as an end-effector of protein regulation?

Publication date: Available online 9 February 2017
Source:Free Radical Biology and Medicine
Author(s): Kathryn Wolhuter, Philip Eaton
Over the last 25 years protein S-nitrosylation, also known more correctly as S-nitrosation, has been progressively implicated in virtually every nitric oxide-regulated process within the cardiovascular system. The current, widely-held paradigm is that S-nitrosylation plays an equivalent role as phosphorylation, providing a stable and controllable post-translational modification that directly regulates end-effector target proteins to elicit biological responses. However, this concept largely ignores the intrinsic instability of the nitrosothiol bond, which rapidly reacts with typically abundant thiol-containing molecules to generate more stable disulfide bonds. These protein disulfides, formed via a nitrosothiol intermediate redox state, are rationally anticipated to be the predominant end-effector modification that mediates functional alterations when cells encounter nitrosative stimuli. In this review we present evidence and explain our reasoning for arriving at this conclusion that may be controversial to some researchers in the field.



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Oxidative DNA Damage & Repair: An Introduction

Publication date: Available online 9 February 2017
Source:Free Radical Biology and Medicine
Author(s): Jean Cadet, Kelvin J.A. Davies
This introductory article should be viewed as a prologue to the Free Radical Biology & Medicine Special Issue devoted to the important topic of Oxidatively Damaged DNA and its Repair. This special issue is dedicated to Professor Tomas Lindahl, co-winner of the 2015 Nobel Prize in Chemistry for his seminal discoveries in the area repair of oxidatively damaged DNA. In the past several years it has become abundantly clear that DNA oxidation is a major consequence of life in an oxygen-rich environment. Concomitantly, survival in the presence of oxygen, with the constant threat of deleterious DNA mutations and deletions, has largely been made possible through the evolution of a vast array of DNA repair enzymes. The articles in this Oxidatively Damaged DNA & Repair special issue detail the reactions by which intracellular DNA is oxidatively damaged, and the enzymatic reactions and pathways by which living organisms survive such assaults by repair processes.



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