HK1 and HK2 Beyond Glycolysis: Mitochondrial Interactions and Dual Roles in Metabolism and Cell Fate.

Gene Therapy Strategies to Restore ER Proteostasis in Disease.

ADAR1-Dependent RNA Editing Promotes MET and iPSC Reprogramming by Alleviating ER Stress.

Identifying critical regulatory interactions in cell fate decision and transition by systematic perturbation analysis

Cells respond actively to the environmental changes, constantly adjusting their structure and function in order to maintain cellular homeostasis. Such changes consequent to both physiological stresses and/or pathological stimuli may be modulated in terms of type, degree, and duration and induce multiple cellular responses. Several different signaling pathways can in fact lead to repair, adaptation, or, in some cases, cell death. A prolonged inflammatory response may ultimately lead to several pathological conditions, including cancer. The specific cell response under stressful conditions has therefore crucial effects not only on the single cell fate but also more generally on tissue homeostasis and ultimately on organism health. This special issue, analyzing different experimental models, aims at highlighting some of the key mechanisms involved in cellular stress response. Topics of this issue include different stressful conditions such as oxidative, hypoxic, endoplasmic reticulum (ER) and inflammatory stress and consequent cell responses such as apoptosis, necroptosis, cell cycle arrest, mitotic cell death, pyroptosis, and survival. Among different types of stress, oxidative stress is causally implicated in degenerative diseases onset. In particular nitrooxidative stress has been reported to be implicated in the loss of specific neuronal populations in Parkinson's disease and amyotrophic lateral sclerosis. In the present special issue, the manuscript entitled “S-Nitrosoglutathione Reductase Plays Opposite Roles in SH-SY5Y Models of Parkinson's Disease and Amyotrophic Lateral Sclerosis” by S. Rizza and collaborators demonstrates that denitrosylating enzyme S-nitrosoglutathione reductase (GSNOR) modulation has different effects on neuronal viability depending on the applied stimulus and highlights GSNOR as a crucial molecular player of neuronal homeostasis. The review entitled “Redox Signaling in Diabetic Nephropathy: Hypertrophy versus Death Choices in Mesangial Cells and Podocytes” by G. Manda and collaborators emphasizes the role of the oxidative stress in diabetic nephropathy, which may act as trigger, modulator, and linker within the complex network of pathologic events. This review particularly addresses the molecular switches deciding on the renal cells fate in diabetic nephropathy leading to hypertrophy versus death choices in mesangial cells and podocytes. On the other hand, amyotrophic lateral sclerosis is a multifactorial disease caused by motor neuron degeneration and is often associated with mutation of the superoxide dismutase 1 (SOD1) gene (encoding an enzyme that uses copper and zinc to break down toxic, charged oxygen molecules called superoxide radicals), thus affecting SOD1 antioxidant activity. The most studied SOD1 mutation is the G93A substitution. The manuscript entitled “Postmitotic Expression of SOD1G93A Gene Affects the Identity of Myogenic Cells and Inhibits Myoblasts Differentiation” by M. Martini and collaborators demonstrates that oxidative stress associated with SOD1G93A expression in C2C12 myogenic cells inhibits their myogenic program and promotes a fibroadipogenic phenotype. Hypoxic stress plays a key role in heart disease, cancer, stroke, and other causes of mortality. The review entitled “Hypoxia Inducible Factor Pathway and Physiological Adaptation: A Cell Survival Pathway?” by H. Kumar and D.-K. Choi describes the role of the Hypoxia Inducible Factors (HIFs) in the survival pathways activated to regulate cellular adaptation to hypoxia. Hypoxia and other environmental factors such as nutrient deprivation, pH changes, and reduced vascularization contribute instead to endoplasmic reticulum stress (ER stress) and activate the Unfolded Protein Response (UPR) in tumor cells. The review “Cancer Microenvironment and Endoplasmic Reticulum Stress Response” by C. Giampietri and collaborators describes the molecular mechanisms of UPR and their role in promoting cancer cells survival and proliferation. Drug Induced Liver Injury (DILI) represents an adverse drug reaction leading to severe liver damage. Kupffer cells sense hepatic stress and produce cytokines thus stimulating an immune response. In the manuscript entitled “Subtoxic Concentrations of Hepatotoxic Drugs Lead to Kupffer Cell Activation in a Human In Vitro Liver Model: An Approach to Study DILI” V. Kegel and collaborators describe a human Kupffer cell culture model in vitro for the investigation of immune-mediated signaling in hepatic inflammatory stress involved in the pathogenesis of DILI. Long-standing inflammatory stress may also promote tumor development, growth, and progression. In the manuscript entitled “Anti-Inflammatory Effects of a Methanol Extract from the Marine Sponge Geodia cydonium on the Human Breast Cancer MCF-7 Cell Line” S. Costantini and collaborators demonstrate that a methanol extract from this marine sponge reduces levels of VEGF and exerts a dose-dependent anti-inflammatory effect. In the manuscript entitled “Multidrug Resistance Protein-4 Influences Aspirin Toxicity in Human Cell Line,” I. Massimi and collaborators investigate aspirin effect on the efflux transporter protein Multidrug Resistance Protein-4 (MRP4) expression in HEK-293 cells. The authors demonstrate that low nontoxic aspirin dosages may lead to MRP-4 overexpression, thus increasing cellular detoxification of aspirin. Since overexpression of efflux transporters in human cells is a mechanism of resistance to drugs, the manuscript by I. Massimi et al. contributes to a better comprehension of molecular mechanisms adopted by cancer cells for survival in stressful conditions during chemotherapy. Various pathological processes such as ischemic brain injury, myocardial infarction, organ transplantation, and virus replication are accompanied by strong inflammatory stress and necroptosis. The manuscript entitled “Necroptotic Cell Death Signaling and Execution Pathway: Lessons from Knockout Mice” by J. Belizario and collaborators describes recent discoveries regarding necroptosis execution pathway. The authors outline and discuss important phenotypes of knockout mice models that serve to define the role of CASPASE 8, FLIP, and FADD genes and other major components of necroptotic signaling. In parallel, pyroptosis is a recently identified type of regulated cell death associated with inflammatory response and with features distinct from both apoptosis and necroptosis. Its role in cardiovascular diseases is addressed in the manuscript “Looking for Pyroptosis-Modulating miRNAs as a Therapeutic Target for Improving Myocardium Survival” by S. Lee et al. The authors aim at elucidating the possible role of miRNAs as therapeutic targets for preventing excessive myocardial pyroptosis in cardiovascular diseases. Progression through the cell cycle is one of the most important decisions during the life of a cell and several kinds of stress are able to influence this choice. The manuscript “Cellular Response upon Stress: p57 Contribution to the Final Outcome” by M. N. Rossi and F. Antonangeli focuses on the contribution of cyclin-dependent kinase inhibitor p57 in regulating cell cycle arrest and apoptosis after cellular stress with particular attention to cancer cells. Finally, the review entitled “Targeting the mitotic catastrophe signaling pathway in cancer” by M. M. Mc Gee describes the molecular mechanism of mitotic cell death, a mechanism to remove defective and genomically unstable cells. Mitotic catastrophe is a regulated antiproliferative process that occurs as a consequence of defective mitosis and may precede apoptosis, thus representing a further response under stressful conditions. The readers will find in this special issue not only interesting data and updated reviews on possible mechanisms of cellular stress response, but also relevant questions waiting to be answered. The study of multiple pathways activated under stressful conditions has become increasingly complex, requiring expertise from all fields of biology. A better comprehension of such pathways might help elucidation of disease pathogenesis and may be of crucial interest in order to develop new therapeutic strategies. Francesco Cecconi Laura Soucek Dennis D. Taub Elio Ziparo

Under conditions of hypoxia, most eukaryotic cells undergo a shift in metabolic strategy, which involves increased flux through the glycolytic pathway. Although this is critical for bioenergetic homeostasis, the underlying mechanisms have remained incompletely understood. Here, we report that the induction of hypoxia-induced glycolysis is retained in cells when gene transcription or protein synthesis are inhibited suggesting the involvement of additional post-translational mechanisms. Post-translational protein modification by the small ubiquitin related modifier-1 (SUMO-1) is induced in hypoxia and mass spectrometric analysis using yeast cells expressing tap-tagged Smt3 (the yeast homolog of mammalian SUMO) revealed hypoxia-dependent modification of a number of key glycolytic enzymes. Overexpression of SUMO-1 in mammalian cancer cells resulted in increased hypoxia-induced glycolysis and resistance to hypoxia-dependent ATP depletion. Supporting this, non-transformed cells also demonstrated increased glucose uptake upon SUMO-1 overexpression. Conversely, cells overexpressing the de-SUMOylating enzyme SENP-2 failed to demonstrate hypoxia-induced glycolysis. SUMO-1 overexpressing cells demonstrated focal clustering of glycolytic enzymes in response to hypoxia leading us to hypothesize a role for SUMOylation in promoting spatial re-organization of the glycolytic pathway. In summary, we hypothesize that SUMO modification of key metabolic enzymes plays an important role in shifting cellular metabolic strategies toward increased flux through the glycolytic pathway during periods of hypoxic stress.

Modulation of Longevity and Tissue Homeostasis by the Drosophila PGC-1 Homolog

Notch signaling and new therapeutic options in liver disease

Multifaceted functions of post-translational enzyme modifications in the control of plant glycolysis

Multicellular organisms develop a wide variety of highly-specialized cell types. The consistency and robustness of developmental cell fate trajectories suggests that complex gene regulatory networks effectively act as low-dimensional cell fate landscapes. A complementary set of works draws on the theory of dynamical systems to argue that cell fate transitions can be categorized into universal decision-making classes. However, the theory connecting geometric landscapes and decision-making classes to high-dimensional gene expression space is still in its infancy. Here, we introduce a phenomenological model that allows us to identify gene expression signatures of decision-making classes from single-cell RNA-sequencing time-series data. Our model combines low-dimensional gradient-like dynamical systems and high-dimensional Hopfield networks to capture the interplay between cell fate, gene expression, and signaling pathways. We apply our model to the maturation of alveolar cells in mouse lungs to show that the transient appearance of a mixed alveolar type 1/type 2 state suggests the triple cusp decision-making class. We also analyze lineage-tracing data on hematopoetic differentiation and show that bipotent neutrophil-monocyte progenitors likely undergo a heteroclinic flip bifurcation. Our results suggest it is possible to identify universal decision-making classes for cell fate transitions directly from data.

Multicellular organisms develop a wide variety of highly-specialized cell types. The consistency and robustness of developmental cell fate trajectories suggests that complex gene regulatory networks effectively act as low-dimensional cell fate landscapes. A complementary set of works draws on the theory of dynamical systems to argue that cell fate transitions can be categorized into universal decision-making classes. However, the theory connecting geometric landscapes and decision-making classes to high-dimensional gene expression space is still in its infancy. Here, we introduce a phenomenological model that allows us to identify gene expression signatures of decision-making classes from single-cell RNA-sequencing time-series data. Our model combines low-dimensional gradient-like dynamical systems and high-dimensional Hopfield networks to capture the interplay between cell fate, gene expression, and signaling pathways. We apply our model to the maturation of alveolar cells in mouse lungs to show that the transient appearance of a mixed alveolar type 1/type 2 state suggests the triple cusp decision-making class. We also analyze lineage-tracing data on hematopoetic differentiation and show that bipotent neutrophil-monocyte progenitors likely undergo a heteroclinic flip bifurcation. Our results suggest it is possible to identify universal decision-making classes for cell fate transitions directly from data.

Ageing is driven by the inexorable and stochastic accumulation of damage in biomolecules vital for proper cellular function. Although this process is fundamentally haphazard and uncontrollable, senescent decline and ageing is broadly influenced by genetic and extrinsic factors. Numerous gene mutations and treatments have been shown to extend the lifespan of diverse organisms ranging from the unicellular Saccharomyces cerevisiae to primates. It is becoming increasingly apparent that most such interventions ultimately interface with cellular stress response mechanisms, suggesting that longevity is intimately related to the ability of the organism to effectively cope with both intrinsic and extrinsic stress. Here, we survey the molecular mechanisms that link ageing to main stress response pathways, and mediate age-related changes in the effectiveness of the response to stress. We also discuss how each pathway contributes to modulate the ageing process. A better understanding of the dynamics and reciprocal interplay between stress responses and ageing is critical for the development of novel therapeutic strategies that exploit endogenous stress combat pathways against age-associated pathologies.

The respiratory pathways of glycolysis, the tricarboxylic acid (TCA) cycle, and mitochondrial electron transport chain are central features of carbon metabolism and bioenergetics in eukaryotic cells. As respiration forms the core of intermediary metabolism, it plays a pivotal role in the growth and metabolism of all photosynthetic organisms. The aim of this chapter is to provide an overview of the occurrence and functions of enzyme post-translational modifications (PTMs) in the control of plant respiration including sucrose catabolism. PTMs are covalent alterations of amino acid residues within a particular polypeptide. Diverse PTMs represent pivotal regulatory events that integrate signaling, gene expression, and metabolism with developmental and stress responses in eukaryotic cells. These PTMs are often rapid and reversible and can not only dramatically alter enzyme activity, but may also generate specific PTM-dependent docking sites that influence interactions with other proteins. In yeast, enzyme PTMs exert more control over glycolysis and mitochondrial metabolism than do changes in transcripts or enzyme abundance, and the same situation likely applies to vascular plants. Recent advances in proteomics, particularly the development of novel and specific chemistries along with affiliated mass spectrometry techniques for detection and mapping of diverse PTMs, are rapidly expanding the catalogue of respiratory enzymes whose functions may be controlled by reversible covalent modification. Phosphorylation-dephosphorylation and disulfide-dithiol interconversion appear to be the most prevalent types of reversible covalent modification used in plant enzyme control. Additional PTMs such as monoubiquitination, S-nitrosylation, and acetylation also appear to play important roles. However, the biochemical impact of in vivo PTMs on the functional properties of plant respiratory enzymes remains mostly unknown and thus remains an important goal for future research. Rational manipulation of PTM events is expected to make an important contribution to the implementation of effective biotechnological strategies for engineering grain crops for increased yields via metabolic engineering.

The Wistar Audiogenic Rat (WAR) strain is a genetic model of epilepsy, specifically brainstem-dependent tonic-clonic seizures, triggered by acute auditory stimulation. Chronic audiogenic seizures (audiogenic kindling) mimic temporal lobe epilepsy, with significant participation of the hippocampus, amygdala, and cortex. The objective of the present study was to characterize the mitochondrial energy metabolism in hippocampus and cortex of WAR and verify its relationship with seizure severity. Hippocampus of WAR naïve (no seizures) presented higher oxygen consumption in respiratory states related to the maximum capacities of phosphorylation and electron transfer system, elevated mitochondrial density, lower GSH/GSSG and catalase activity, and higher protein carbonyl and lactate contents, compared with their Wistar counterparts. Audiogenic kindling had no adding functional effect in WAR, but in Wistar, it induced the same alterations observed in the audiogenic strain. In the cortex, WAR naïve presented elevated mitochondrial density, lower GSH/GSSG and catalase activity, and higher protein carbonyl levels. Chronic acoustic stimulation in Wistar induced the same alterations in cortex and hippocampus. Mainly in the hippocampus, WAR naïve presented elevated mRNA expression of glucose, lactate and excitatory amino acids transporters, several glycolytic enzymes, lactate dehydrogenase, and Na+/K+ ATPase in neurons and in astrocytes. In vivo treatment with mitochondrial uncoupler 2,4-dinitrophenol (DNP) or N-acetylcysteine (NAC) in WAR had no effect on mitochondrial metabolism, but lowered oxidative stress. Unlike DNP, NAC downregulated all enzyme genes involved in glucose and lactate uptake, and metabolism in neurons and astrocytes. Additionally, it was able to reduce brainstem seizure severity in WAR. In conclusion, in WAR naïve animals, both cerebral cortex and hippocampus display elevated mitochondrial density and/or activity associated with oxidative damage, glucose and lactate metabolism pathways upregulation, and increased Na+/K+ ATPase mRNA expression. Only in vivo treatment with NAC was able to reduce seizure severity of kindled WARs, possibly via down regulation of glucose/lactate metabolism. Taken together, our results are a clear contribution to the field of mitochondrial metabolism associated to epileptic seizures.

Cell fate transitions are common in many developmental processes. Therefore, identifying the mechanisms behind them is crucial. Traditionally, due to complexity of networks and existence of plenty of kinetic parameters, dynamical analysis of biomolecular networks can only be performed by simultaneously perturbing a small number of parameters. Although many efforts have focused on how cell states change under specific perturbations, conversely, how to infer parametric conditions underlying distinct cell fates by systematic perturbations is less clear and needs to be further investigated. In this article, we present a general computational method by integrating systematic perturbations, unsupervised clustering, principal component analysis, and fitting analysis. The method can be used to to construct maps between distinct cell fates and parametric conditions by systematic perturbations. In particular, there are no needs of accurate parameter measurements and occurrence of bifurcations to establish the maps. To validate feasibility and inference performance of the method, we use toggle switch, inner cell mass, and epithelial mesenchymal transition as model systems to show how the maps are constructed and how system parameters encode essential information on cell fates. The maps tell us how systematic perturbations drive cell fate decisions and transitions, and allow us to purposefully predict, manipulate, and even control cell states. The approach is especially helpful in understanding crucial roles of certain parameter combinations during fate transitions. We hope that the approach can provide us valuable information on parametric or perturbation conditions so some specific targets, e.g. directional differentiation, can be realized. No public data are used. The data we used are generated by randomly chosen values of model parameters in certain ranges, and the corresponding parameters are already attached in supplementary materials.

Restraining Lysosomal Activity Preserves Hematopoietic Stem Cell Quiescence and Potency.