Kinases and Phosphatases

Non-tuberculous mycobacteria (NTM) represent a diverse group of mycobacterial species known for causing opportunistic infections, especially in individuals with underlying health conditions. Unlike Mycobacterium tuberculosis (Mtb), the causative agent of tuberculosis, NTM species exhibit different pathogenic characteristics and drug resistance mechanisms, making them increasingly relevant in clinical settings. PtkA is a crucial protein tyrosine kinase that regulates bacterial growth, stress response, and virulence by phosphorylating various substrates in Mtb. Understanding whether PtkA homologs exist in NTM could provide insights into their virulence and resistance mechanisms. In silico approaches, which utilize computational tools for sequence alignment, structure prediction, and functional annotation, offer a powerful means to identify homologous proteins across different species. In this article, we have employed tools like BLAST (Basic Local Alignment Search Tool), protein structure databases, and the NTM database to identify PtkA homologs in NTM genomes, providing a foundation for further studies. Full article

Protein kinases and phosphatases are key signaling proteins and are important drug targets. An explosion in the number of publicly available 3D structures of proteins has been seen in recent years. Three-dimensional structures of kinase and phosphatase have not been systematically investigated. This is due to the difficulty of designing structure-based descriptors that are capable of quantifying conformational changes. We have developed a triangular spatial relationship (TSR)-based algorithm that enables a unique representation of a protein’s 3D structure using a vector of integers (keys). The main objective of this study is to provide structural insight into conformational changes. We also aim to link TSR-based structural descriptors to their functions. The 3D structures of 2527 kinases and 505 phosphatases are studied. This study results in several major findings as follows: (i) The clustering method yields functionally coherent clusters of kinase and phosphatase families and their superfamilies. (ii) Specific TSR keys are identified as structural signatures for different types of kinases and phosphatases. (iii) TSR keys can identify different conformations of the well-known DFG motif of kinases. (iv) A significant number of phosphatases have their own distinct DFG motifs. The TSR keys from kinases and phosphatases agree with each other. TSR keys are successfully used to represent and quantify conformational changes of CDK2 upon the binding of cyclin or phosphorylation. TSR keys are effective when used as features for unsupervised machine learning and for key searches. If discriminative TSR keys are identified, they can be mapped back to atomic details within the amino acids involved. In conclusion, this study presents an advanced computational methodology with significant advantages in not only representing and quantifying conformational changes of protein structures but also having the capability of directly linking protein structures to their functions. Full article

Chemical studies usually consist of measurements made on large ensembles of molecules with data representing average values for the population. It has been shown that individual molecules of a given enzyme have different properties. Large-scale averaging has in the past masked these differences. Alkaline phosphatase has been used as a model to study this enzyme heterogeneity. The catalytic rates of the individual molecules have been found to differ by over 10-fold, and the activation energy of catalysis by more than two-fold. Differences in properties indicate that differences in structure must exist between the molecules. For alkaline phosphatase, the structural differences have been suggested to be differences in glycosylation, differences due to partial proteolysis, and due to some molecules containing mixtures of active and inactive subunits. The determination of the distribution of activities of populations of this enzyme within a sample has also been shown to be a useful tool in diagnostics. This review discusses the advent of single-molecule enzymology and summarizes its use in the study of alkaline phosphatase using capillary electrophoresis, microscopic well assays, and single-molecule tracking. Full article

Angiogenesis is essential for remodeling and repairing existing vessels, and this process requires signaling pathways including those controlled by transforming growth factor beta (TGF-β). We have previously reported crosstalk between TGF-β and the protein kinase With No lysine (K) 1 (WNK1). Homozygous disruption of the gene encoding WNK1 results in lethality in mice near embryonic day E12 due to impaired angiogenesis, and this defect can be rescued by the endothelial-specific expression of an activated form of the WNK1 substrate kinase Oxidative Stress-Responsive 1 (OSR1). However, molecular processes regulated via a collaboration between TGF-β and WNK1/OSR1 are not well understood. Here, we show that WNK1 interacts with the E3 ubiquitin ligases SMURF1/2. In addition, we discovered that WNK1 regulates SMURF1/2 protein stability and vice versa. We also demonstrate that WNK1 activity regulates TGF-β receptor levels, in turn, controlling TGF-β signaling. Full article

Obesity has become a global epidemic, contributing to various metabolic diseases. Despite existing therapies, the need to investigate new molecular targets to combat obesity-associated pathologies persists. Glycogen Synthase Kinase-3 (GSK-3), a serine/threonine kinase with two paralogs (GSK-3α and GSK-3β), has emerged as a critical player in obesity-associated metabolic pathologies such as type 2 diabetes (T2D), and cardiovascular diseases (CVDs). However, its ubiquitous dynamic expression and complex context-dependent signaling pathways present challenges in understanding its precise role in metabolic perturbations. In the present review, we will highlight the specific role and the proposed mechanisms via which the two GSK-3 paralogs impact obesity-associated pathologies such as T2D, diabetic cardiomyopathy (DCM), and cognitive impairment, a hallmark of Alzheimer’s disease (AD). We will also highlight studies delineating the role of GSK-3s using either GSK-3 inhibitors or non-pharmacological compounds to inhibit/taper GSK-3 activity in metabolic diseases. Thus, the primary goal of this review is to highlight recent findings delineating the regulation/dysregulation of GSK-3α/β in tissues such as heart, liver, skeletal muscle, pancreas, brain, and adipose tissue that undergo morphological and metabolic changes with diet-induced obesity which predisposes obese individuals to numerous devastating chronic conditions by GSK-3 overactivity. Full article

ATM kinase is becoming an important therapeutic target for tumor radiosensitization. Radiation is known to cause neuro-inflammation and neurodegeneration; however, the effects of small molecule ATM inhibitors (ATMi’s) and radiation on normal tissue, including healthy brain, are largely unexplored. Therefore, we examined the mouse CNS after ATMi radiosensitization with a focus on the fate of neurons. We used several approaches to assess the effects on the DNA damage response (DDR) and apoptosis of neurons using immunostaining. In vivo, a significant decrease in viable neurons and increase in degenerating neurons and apoptosis was observed in mice treated with radiation alone. On the other hand, an ATMi alone had little to no effect on neuron viability and did not induce apoptosis. Importantly, the ATMi’s did not further increase radiation toxicity. In fact, multiplex immunostaining showed that a clinical candidate ATMi (AZD1390) protected mouse neurons from apoptosis by 90% at 4 h after radiation. We speculate that the lack of toxicity to neurons is due to a normal ATM–p53 response that, if blocked transiently with an ATMi, is protective. Altogether, in line with previous work using ATM knockout mice, we provide evidence that ATM kinase inhibition using small molecules does not add to neuronal radiation toxicity, and might, in fact, protect them from radiation-induced apoptosis at least in the short term. Full article

Cutaneous T-cell lymphomas (CTCLs) are a heterogeneous group of T-cell lymphomas characterised by high relapse rates and no curative treatments unless the allogeneic stem cell transplantation. The main complication in the management of this kind of malignancy is the variability that characterises the genetic and clinical features among the CTCL subtypes. JAK/STAT, MAPK/ERK, PI3K/Akt, and NF-kB are those signalling pathways that are found altered in CTCL and that are responsible for promoting both T-cell malignancy and the pro-tumorigenic microenvironment. Thus, targeting key players of these pathways can be an advantageous therapeutic option for CTCL. In this review, we aim to summarise the different approaches that precisely inhibit the kinases of each cited signalling. JAK inhibitors seem to be the most promising kinase inhibitors for CTCL. However, adverse events have been reported especially in patients with immunosuppression or an underlying autoimmune disease. More studies are needed, especially clinical trials, to investigate the benefits of these drugs for the treatment of cutaneous T-cell lymphomas. Full article

Copper is an essential heavy metal for diverse biological functions but toxic in excess. Consequently, a tightly regulated protein system is required to ensure adequate intracellular levels. In recent decades, several studies have explored the role of Cu⁺-ATPases in copper transport and homeostasis, revealing that these proteins are subject to kinase-mediated phosphorylation that significantly impacts their function. Techniques such as phosphoproteomic screening, site-directed mutagenesis, and artificial neural network tools demonstrated the regulatory effect of phosphorylation on these ATPases. Different protein kinases regulate Cu⁺-ATPases, modulating the active copper transport by affecting specific steps of the catalytic cycle, long-range intramolecular crosstalks, protein trafficking, gene expression, and protein stability. Therefore, the regulatory phosphorylation of Cu⁺-ATPases by kinases ultimately influences the intracellular copper distribution. This study aims to present a review of the scientific literature on the regulation of Cu⁺-ATPases by kinase-mediated phosphorylation as a crucial mechanism for copper homeostasis. This regulation offers new perspectives for developing therapies for disorders related to copper metabolism, such as Wilson and Menkes diseases, as well as cancer, diabetes mellitus, Parkinson’s, and Alzheimer’s diseases. These findings emphasize the need to further comprehend the signaling pathways involving protein kinases in the context of copper regulation. Full article

The phosphorylation state of 20 types of intracellular proteins in the presence of the protein phosphatase 1 (PP1)- and PP2A-specific Ser/Thr phosphatase inhibitor calyculin A or the Tyr phosphatase inhibitor pervanadate was visualized by Phos-tag SDS-PAGE followed by immunoblotting. All blots showed a Phos-tag pattern indicating increased phosphorylation in the presence of one or both phosphatase inhibitors. The increase in phosphorylation stoichiometry per protein tends to be greater for Ser/Thr phosphatase inhibition than for Tyr phosphatase inhibition. This is consistent with the fact that the number of Ser/Thr kinase genes in the human genome is greater than that of Tyr kinases and with the fact that the phospho-Ser/phospho-Thr ratio in the actual human phosphoproteome is far greater than that of phospho-Tyr ratio. This suggests that cellular proteins are routinely and randomly phosphorylated by different kinases with no biological significance, simply depending on the frequency of substrate encounters. Phosphatase is responsible for routinely removing these unwanted phosphate groups systematically and maintaining the dynamic equilibrium of physiological protein phosphorylation. Phos-tag SDS-PAGE visualized that the kinase reaction involves many incidental phosphorylation and that phosphatases play broader roles besides being strict counterparts to kinases. Full article

Protein phosphorylation serves as a fundamental regulatory mechanism to modulate cellular responses to environmental stimuli and plays a crucial role in orchestrating adaptation and metabolic homeostasis in various diverse organisms. In cyanobacteria, an ancient phylum of significant ecological and biotechnological relevance, protein phosphorylation emerges as a central regulatory axis mediating adaptive responses that are essential for survival and growth. This exhaustive review thoroughly explores the complex terrain of protein phosphorylation in cyanobacterial adaptation and metabolism, illustrating its diverse forms and functional implications. Commencing with an overview of cyanobacterial physiology and the historical trajectory of protein phosphorylation research in prokaryotes, this review navigates through the complex mechanisms of two-component sensory systems and their interplay with protein phosphorylation. Furthermore, it investigates the different feeding modes of cyanobacteria and highlights the complex interplay between photoautotrophy, environmental variables, and susceptibility to photo-inhibition. The significant elucidation of the regulatory role of protein phosphorylation in coordinating light harvesting with the acquisition of inorganic nutrients underscores its fundamental importance in the cyanobacterial physiology. This review highlights its novelty by synthesizing existing knowledge and proposing future research trajectories, thereby contributing to the deeper elucidation of cyanobacterial adaptation and metabolic regulation through protein phosphorylation. Full article

Mitochondria are organelles involved in cellular energetics in all eukaryotes, and changes in their dynamics, fission, fusion, or localization can lead to cell defects and disease in humans. Budding yeast, Saccharomyces cerevisiae, has been shown to be an effective model organism in elucidating mechanisms underpinning these mitochondrial processes. In the work presented here, a genetic screen was performed to identify overexpressing kinases, phosphatases, and ubiquitin ligases, which resulted in mitochondrial defects. A total of 33 overexpressed genes showed mitochondrial phenotypes but without severe growth defects. These included a subset that affected the timing of mitochondrial inheritance and were the focus of further study. Using cell and biochemical approaches, the roles of the PAK-family kinase Cla4 and the E3-ubiquitin ligases Dma1 and Dma2 were investigated. Previous studies have indicated the roles of kinase Cla4 and ligases Dma1 and Dma2 in triggering the degradation of trafficking adaptors in the bud, which leads to disruption of the interaction with the transporting class V myosin, Myo2. Here, we map a key interface between Cla4 and the mitochondrial adaptor Mmr1 necessary for phosphorylation and identify a region of Mmr1 required for its degradation via Dma1 and Dma2. Together, our data provide insights into key regulatory regions of Mmr1 responsible for its function in mitochondrial inheritance. Full article

Cancer, characterized by uncontrolled cell growth and metastasis, represents a significant challenge to public health. The IGF1/IGF1R axis plays a pivotal role in tumor proliferation and survival, presenting an attractive target for intervention. NT157, a small molecule tyrphostin, has emerged as a promising inhibitor of this axis, displaying potent antineoplastic effects across various cancer types. This review synthesizes the literature on NT157’s mechanism of action and its impact on cellular processes in experimental cancer models. Initially identified for inducing the serine phosphorylation of IRS1 and IRS2, leading to their degradation and inhibiting the IGF1R signaling cascade, subsequent studies revealed additional targets of NT157, including STAT3, STAT5, and AXL, suggesting a multifaceted mechanism. Experimental evidence demonstrates that NT157 effectively suppresses tumor growth, metastasis, and angiogenesis in diverse cancer models. Additionally, NT157 enhances chemotherapy efficacy in combination therapy. Moreover, NT157 impacts not only tumor cells but also the tumor microenvironment, modulating inflammation and immune responses by targeting cancer-associated fibroblasts, myeloid cells, and immune cells, creating a suppressive milieu hindering tumor progression and metastasis. In conclusion, NT157 exhibits remarkable versatility in targeting multiple oncogenic pathways and hallmarks of cancer, underscoring its potential as a promising therapeutic agent. Full article

It has been proposed that procedures which upregulate mitochondrial biogenesis and autophagy by replacing damaged mitochondria with healthy ones may prevent the development of several heart diseases. A member of serine and threonine kinases, adenosine monophosphate-activated protein kinase (AMPK), could play essential roles in the autophagy and/or mitophagy. AMPK is widely distributed in various cells, which might play diverse regulatory roles in different tissues and/or organs. In fact, changes in the kinase function of AMPK due to alteration of activity have been linked with diverse pathologies including cardiac disorders. AMPK can regulate mitochondrial biogenesis via peroxisome proliferator-activated receptor gamma coactivator 1-alpha (PGC1α) signaling and also improve oxidative mitochondrial metabolism through inhibition of mechanistic/mammalian target of rapamycin (mTOR) pathway, which may also modulate the autophagy/mitophagy through autophagy activating kinase 1 (ULK1) and/or transforming growth factor beta (TGF-β) signaling. Therefore, the modulation of AMPK in autophagy/mitophagy pathway might probably be thought as a therapeutic tactic for several cardiac disorders. As kinases are amongst the most controllable proteins, in general, the design of small molecules targeting kinases might be an eye-catching avenue to modulate cardiac function. Some analyses of the molecular biology underlying mitophagy suggest that nutraceuticals and/or drugs including specific AMPK modulator as well as physical exercise and/or dietary restriction that could modulate AMPK may be useful against several heart diseases. These observations may virtually be limited to preclinical studies. Come to think of these, however, it is speculated that some nutraceutical regimens might have positive potential for managing some of cardiac disorders. Full article

Cancer stem cells (CSCs), found within tumors, are powerful drivers of disease recurrence and metastasis. Their abilities to self-renew and maintain stem-like properties make treatment difficult, as their heterogeneity and metastatic properties can lead to resistance and limit the effectiveness of standard therapies. Given their significance, CSCs are typically isolated based on combinations of markers, which often indicate heterogeneous populations of CSCs. The lack of consensus in cell characterization poses challenges in defining and targeting these cells for effective therapeutic interventions. In this review, we suggest five promising molecules—ABCB5, CD26, CD66c, uPAR, and Trop-2—chosen specifically for their distinct distribution within cancer types and clinical relevance. These markers, expressed at the cell surface of CSCs, could significantly enhance the specificity of cancer stemness characterization. This review focuses on describing their pivotal roles as biomarker checkpoints for metastasis. Additionally, this review outlines existing literature on glycosylation modifications, which present intriguing epitopes aimed at modulating the stability and function of these markers. Finally, we summarize several promising in vivo and clinical trial approaches targeting the mentioned surface markers, offering potential solutions to overcome the therapeutic resistance of CSCs and addressing current gaps in treatment strategies. Full article

Short-chain fatty acids (SCFAs), derived from fermentation of dietary fibers and resistant starch by the microbiota in the colon, exert multiple effects on colonic functions, including tumor suppressing activities. Our previous studies found that SCFAs induced autophagy in colon cancer cells via downregulating mTOR signaling, but the mechanism involved in mTOR suppression still needs to be defined. In this study, we identified rhabdomyosarcoma 2 associated transcript (RMST), a long non-coding RNA, as a key mediator for SCFAs to suppress mTOR activation in colon cancer cells. RMST could be significantly induced by SCFAs in a time- and dose-dependent manner. RMST, by itself, was sufficient to suppress mTOR signaling and augment autophagosome formation. Depletion of RMST, through siRNA or CRISPR knockdown, reduced the abilities of SCFAs to suppress mTOR activation or to induce autophagic responses. RMST increased the expression level of TSC2, a negative regulator of the mTOR signaling pathway. Our data delineate a novel RMST/TSC2 cellular pathway, enlisted by SCFAs, to modulate mTOR activities in colon cancer cells. Full article

CK2 is a protein kinase that plays an important role in numerous cellular pathways involved in cell growth, differentiation, proliferation, and death. Consequently, upregulation of CK2 is implicated in many disease types, in particular cancer. As such, CK2 has gained significant attention as a potential therapeutic target in cancer, and over 40 chemical probes targeting CK2 have been developed in the past decade. In this review, we highlighted several chemical probes that target sites outside the conventional ATP-binding site. These chemical probes belong to different classes of molecules, from small molecules to peptides, and possess different mechanisms of action. Many of the chemical probes discussed in this review could serve as promising new candidates for drugs selectively targeting CK2. Full article

Kinases play crucial roles in the pathophysiology of retinal degenerative diseases. These diseases, such as diabetic retinopathy, age-related macular degeneration, glaucoma, and retinitis pigmentosa, are characterized by progressive degeneration of retinal cells, including photoreceptors, ganglion cells, vascular cells, and retinal pigment epithelium, among others. The involvement of kinases in cell survival and apoptosis, immune responses and inflammation regulation, mitochondrial functions and mitophagy, autophagy, and proteostasis is crucial for maintaining cellular homeostasis and responding to various stressors. This review highlights the importance of studying kinases to better understand their functions and, regulation permitting, enable the identification of novel molecular players or potential drug targets and, consequently, the development of more effective and precise treatments to slow or halt the progression of retinal degenerative diseases. Full article

Tissue transglutaminase2 (TG2) has emerged as a key enigmatic protein in the development of various metabolic and age-related diseases. It catalyzes covalent cross-linking of countless proteins and provides strength to the extracellular matrix and resistance to proteolytic degradation via different pathways, including NF-kβ, TGF-β and PI3K/Akt as the major signaling pathways. The etiology of diabetes and associated diseases has been found to be linked to unbalanced TG2 activity that may not only result in impaired or delayed wound healing in diabetics but also worsen degenerative and metabolic disease conditions. TG2 is usually overexpressed in diabetes, fibrosis, cancer, and neurodegenerative disorders. These TG2-linked diseases are usually associated with prolonged activation of inflammatory pathways. Therefore, reducing the inflammatory mechanisms and improving tissue remodeling appear to be the main treatment strategies to exterminate TG2-linked diseases. The present review aims to deliver a detailed overview of the existing understanding of TG2 in diabetes and associated diseases’ progression, as well as treatment strategies to regulate TG2 tightly and its potential clinical applications. Our research endorses the notion that TG2 can serve as an effective early-stage diagnostic biomarker for metabolic diseases and a therapeutic target for the development of potential drug. Full article

Protein kinase CK2 (CK2) influences one-fifth of the cellular phosphoproteome. It regulates almost all cellular pathways and is thus a critical switch between biological processes within a cell. Inhibition of CK2 reverses oncogene addiction of tumor and alters tumor microenvironment. The success of this strategy and its clinical translation opens new opportunities. Targeting CK2 in musculoskeletal disorders is promising. Clinical manifestations of these disorders include dysfunctional inflammation, dysregulated cell differentiation, and senescence. Processes regulated by CK2 include all of these. Its emerging role in senescence also indicates its function’s centrality in cellular metabolism. This review summarizes considerations for targeting CK2 in musculoskeletal disorders. We have discussed the implications of CK2-regulated processes in musculoskeletal disorders. Full article