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Salinity stress is a major concern in regions where irrigation relies on saline water. This study aimed to investigate the relative water content (RWC), electrolytic leakage (EL), total chlorophyll content, free amino acid content, and total soluble sugar content were analyzed in different groundnut species subjected to various salinity treatments. The results showed that salinity stress significantly reduced the RWC in groundnut leaves, with A. duranensis (wild type) exhibiting higher RWC values compared to the Arachis hypogaea species. RNA sequencing was performed to identify differentially expressed genes (DEGs) during salt stress. A total of 9079 DEGs were identified, with 1372 genes upregulated and 2509 genes downregulated. Genes belonging to transcription factor families, such as WRKY, MYB, bHLH, E2F, and Auxin efflux carrier proteins, were induced under salt stress in the tolerant genotype. Conversely, genes encoding NADH dehydrogenase, glutathione S-transferase, protein kinases, UDP-glycosyltransferase, and peroxidase were downregulated. Gene ontology and pathway analyses revealed several enriched categories and metabolic pathways associated with salt stress response, including catalytic activity, response to salt stress, ATP-dependent activity, and oxidative phosphorylation. The findings of this study provide insights into the physiological and molecular responses of groundnut to salinity stress. A. duranensis exhibited better salinity tolerance than Arachis hypogaea , as indicated by higher RWC values, lower electrolytic leakage, and differential gene expression patterns. These results contribute to our understanding of the mechanisms underlying salt stress tolerance in groundnut and may guide future efforts to develop salinity-tolerant groundnut species, ultimately improving crop yield in saline-affected regions. • Salinity stress decreases crop yield by affecting water content and leaf damage in groundnut plants. • A. duranensis variety shows higher tolerance to salinity stress compared to GG20. • Salinity stress reduces chlorophyll levels in groundnut leaves, with A. duranensis being more resilient. • Salinity stress triggers the accumulation of free amino acids and soluble sugars, especially in A. duranensis. • RNA-seq analysis reveals key genes involved in groundnut's response to salt stress, providing valuable insights. [ABSTRACT FROM AUTHOR]

The phosphoproteome is vast and intricate, with many aspects of its extent and site-specific functional roles still largely unexplored. While traditional studies have primarily focused on Ser/Thr and Tyr phosphorylation, new methodologies are emerging to study non-canonical phosphorylation events. Protein phosphatases play critical roles in cellular signaling, with recent studies highlighting their complexity and involvement in various biological processes. A significant portion of the kinome, referred to as the 'dark' kinome, remains understudied and holds great potential for novel discoveries. Pseudokinases and pseudophosphatases, despite lacking catalytic activity, play crucial roles in cell signaling. Recent advances in experimental and computational analyses have improved the understanding of kinase–substrate and phosphatase–substrate interactions. Protein phosphorylation is a crucial regulatory mechanism in cell signaling, acting as a molecular switch that modulates protein function. Catalyzed by protein kinases and reversed by phosphoprotein phosphatases, it is essential in both normal physiological and pathological states. Recent advances have uncovered a vast and intricate landscape of protein phosphorylation that include histidine phosphorylation and more unconventional events, such as pyrophosphorylation and polyphosphorylation. Many questions remain about the true size of the phosphoproteome and, more importantly, its site-specific functional relevance. The involvement of unconventional actors such as pseudokinases and pseudophosphatases adds further complexity to be resolved. This review explores recent discoveries and ongoing challenges, highlighting the need for continued research to fully elucidate the roles and regulation of protein phosphorylation. [ABSTRACT FROM AUTHOR]

After ejaculation, mammalian sperm undergo a series of molecular events conducive to the acquisition of fertilizing competence. These events are collectively known as capacitation and involve acrosomal responsiveness and a vigorous sperm motility called hyperactivation. When mimicked in the laboratory, capacitating bovine sperm medium contains bicarbonate, calcium, albumin and heparin, among other components. In this study, we aimed at establishing a new capacitation protocol for bovine sperm, using calcium ionophore. Similar to our findings using mouse sperm, bovine sperm treated with Ca2+ ionophore A23187 were quickly immobilized. However, these sperm initiated capacitation after ionophore removal in fresh medium without heparin, and independent of the Protein Kinase A. When A23187-treated sperm were used on in vitro fertilization (IVF) procedures without heparin, eggs showed cleavage rates similar to standardized IVF protocols using heparin containg synthetic oviduct fluid (IVF-SOF). However, when A23187 pre-treated sperm were further used for inseminating eggs in complete IVF-SOF-heparin, a significantly higher percentage of embryo development was observed, suggesting a synergism between two different signaling pathways during bovine sperm capacitation. These results have the potential to improve current protocols for bovine IVF that could also be applied in other species of commercial interest. • Exposure calcium ionophore improves bovine sperm hyperactivation after washing. • Sperm capacitation is achieved by transient exposure to calcium ionophore. • Exposure calcium ionophore trigger capacitation events independent of PKA. • Calcium ionophore capacitated sperm improve embryo production when used in IVF. [ABSTRACT FROM AUTHOR]

Endothelial cAMP-specific phosphodiesterase PDE3A is one of the major negative regulators of the endothelial barrier function in acute lung injury models. However, the mechanisms underlying its regulation still need to be fully resolved. We show here that the PDE3A is a newly described client of the molecular chaperone heat shock protein 90 (hsp90). In endothelial cells (ECs), hsp90 inhibition by geldanamycin (GA) led to a disruption of the hsp90/PDE3A complex, followed by a significant decrease in PDE3A protein levels. The decrease in PDE3A protein levels was ubiquitin-proteasome-dependent and required the activity of the E3 ubiquitin ligase C terminus of Hsc70-interacting protein. GA treatment also enhanced the association of PDE3A with hsp70, which partially prevented PDE3A degradation. GA-induced decreases in PDE3A protein levels correlated with decreased PDE3 activity and increased cAMP levels in EC. We also demonstrated that protein kinase G-dependent phosphorylation of PDE3A at Ser654 can signal the dissociation of PDE3A from hsp90 and PDE3A degradation. This was confirmed by endogenous PDE3A phosphorylation and degradation in 8-Br-cGMP-or 8-CPT-cGMP- and Bay 41-8543-stimulated EC and comparisons of WT- and phospho-mimic S654D mutant PDE3A protein stability in transiently transfected HEK293 cells. In conclusion, we have identified a new mechanism of PDE3A regulation mediated by the ubiquitin-proteasome system. Further, the degradation of PDE3A is controlled by the phosphorylation of S654 and the interaction with hsp90. We speculate that targeting the PDE3A/hsp90 complex could be a therapeutic approach for acute lung injury. [ABSTRACT FROM AUTHOR]

Ubiquitin-specific proteases (USPs) are a family of multidomain deubiquitinases (DUBs) with variable architectures, some containing regulatory auxiliary domains. Among the USP family, all occurrences of intramolecular regulation presently known are autoactivating. USP8 remains the sole exception as its putative WW-like domain, conserved only in vertebrate orthologs, is autoinhibitory. Here, we present a comprehensive structure-function analysis describing the autoinhibition of USP8 and provide evidence of the physical interaction between the WW-like and catalytic domains. The solution structure of full-length USP8 reveals an extended, monomeric conformation. Coupled with DUB assays, the WW-like domain is confirmed to be the minimal autoinhibitory unit. Strikingly, autoinhibition is only observed with the WW-like domain in cis and depends on the length of the linker tethering it to the catalytic domain. Modeling of the WW:CD complex structure and mutagenesis of interface residues suggests a novel binding site in the S1 pocket. To investigate the interplay between phosphorylation and USP8 autoinhibition, we identify AMPactivated protein kinase as a highly selective modifier of S718 in the 14-3-3 binding motif. We show that 14-3-3g binding to phosphorylated USP8 potentiates autoinhibition in a WW-like domain-dependent manner by stabilizing an autoinhibited conformation. These findings provide mechanistic details on the autoregulation of USP8 and shed light on its evolutionary significance. [ABSTRACT FROM AUTHOR]

Mutations in LRRK2 are the most common causes of Parkinson's disease (PD). Dysfunction in LRRK2 enzymatic activities and elevated protein levels are associated with the disease. Higher-order dimerization/oligomerization and membrane recruitment appear to be required for LRRK2 activation. Rab29/12/38/32/8/10 and VPS35 are emerging upstream activators of LRRK2. A unique feature of LRRK2 is that it possesses two enzymatic functions: kinase and GTPase. It phosphorylates a group of proteins including Rabs and its autophosphorylation site S1292. LRRK2 GTPase regulation and contribution to PD is understudied. LRRK2 plays a diverse role in multiple cellular pathways. LRRK2 could be targeted on enzymatic activities, protein levels, crucial cellular interactors, and dimerization. Targeting LRRK2 kinase activity and protein levels has shown promise, and some approaches are advancing to clinical trials. Mutations in leucine-rich repeat kinase 2 (LRRK2) are the most common causes of Parkinson's disease (PD) to date. Dysfunction in LRRK2 enzymatic activities and elevated protein levels are associated with the disease. How is LRRK2 activated, and what downstream molecular and cellular processes does LRRK2 regulate? Addressing these questions is crucial to decipher the disease mechanisms. In this review we focus on the upstream regulations and briefly discuss downstream substrates of LRRK2 as well as the cellular consequences caused by these regulations. Building on these basic findings, we discuss therapeutic strategies targeting LRRK2 and highlight the challenges in clinical trials. We further highlight the important questions that remains to be answered in the LRRK2 field. [ABSTRACT FROM AUTHOR]

Sugar signaling governed by SNF1-RELATED PROTEIN KINASE 1 (SnRK1), TARGET OF RAPAMYCIN (TOR), and trehalose 6-phosphate (T6P) balances energy for root growth. T6P emerges as a central regulator linking auxin, SnRK1, and TOR in root development. A mechanism is emerging in which SnRK1 inhibits TOR through phosphorylation, while T6P can obstruct SnRK1 activity through interaction with TPS class II proteins, thereby controlling the energy that can be used for root development. Sugars derived from photosynthesis, specifically sucrose, are the primary source of plant energy. Sucrose is produced in leaves and transported to the roots through the phloem, serving as a vital energy source. Environmental conditions can result in higher or lower photosynthesis, promoting anabolism or catabolism, respectively, thereby influencing the sucrose budget available for roots. Plants can adjust their root system to optimize the search for soil resources and to ensure the plant's adaptability to diverse environmental conditions. Recently, emerging research indicates that SNF1-RELATED PROTEIN KINASE 1 (SnRK1), trehalose 6-phosphate (T6P), and TARGET OF RAPAMYCIN (TOR) collectively serve as fundamental regulators of root development, together forming a signaling module to interpret the nutritional status of the plant and translate this to growth adjustments in the below ground parts. [ABSTRACT FROM AUTHOR]

Extracellular signal-regulated protein kinase 5 (Erk5), a member of the mitogen-activated protein kinase (MAPK) family, is ubiquitously expressed in all eukaryotic cells and is implicated in the various mitotic processes such as cell survival, proliferation, migration, and differentiation. However, the potential functional roles of Erk5 in oocyte meiosis have not been fully determined. In this study, we document that ERK5 participates in the meiotic maturation of mouse oocytes by regulating the spindle assembly to ensure the meiotic progression. We unexpectedly found that phosphorylated ERK5 was localized in the spindle pole region at metaphase I and II stages by immunostaining analysis. Inhibition of ERK5 activity using its specific inhibitor XMD8-92 dramatically reduced the incidence of first polar body extrusion. In addition, inhibition of ERK5 evoked the spindle assembly checkpoint to arrest oocytes at metaphase I stage by impairing the spindle assembly, chromosome alignment and kinetochore-microtubule attachment. Mechanically, over-strengthened microtubule stability was shown to disrupt the microtubule dynamics and thus compromise the spindle assembly in ERK5-inhibited oocytes. Conversely, overexpression of ERK5 caused decreased level of acetylated α-tubulin and spindle defects. Collectively, we conclude that ERK5 plays an important role in the oocyte meiotic maturation by regulating microtubule dynamics and spindle assembly. • Phosphorylated ERK5 was localized in the spindle pole region at metaphase I and II stages in oocytes. • Inhibition of ERK5 evoked the spindle assembly checkpoint to arrest oocytes at metaphase I stage. • Over-strengthened microtubule stability was shown to disrupt the microtubule dynamics in ERK5-inhibited oocytes. • Overexpression of ERK5 causes decreased level of acetylated α-tubulin and spindle defects. [ABSTRACT FROM AUTHOR]

An ever-growing number of studies highlight the importance of S-acylation, a reversible protein-lipid modification, for diverse aspects of intracellular signaling. In this review, we summarize the current understanding of how S-acylation regulates perhaps the best-known class of signaling enzymes, protein kinases. We describe how S-acylation acts as a membrane targeting signal that localizes certain kinases to specific membranes, and how such membrane localization in turn facilitates the assembly of signaling hubs consisting of an S-acylated kinase’s upstream activators and/or downstream targets. We further discuss recent findings that S-acylation can control additional aspects of the function of certain kinases, including their interactions and, surprisingly, their activity, and how such regulation might be exploited for potential therapeutic gain. We go on to describe the roles and regulation of de-S-acylases and how extracellular signals drive dynamic (de) S-acylation of certain kinases. We discuss how S-acylation has the potential to lead to “emergent properties” that alter the temporal profile and/or salience of intracellular signaling events. We close by giving examples of other S-acylation– dependent classes of signaling enzymes and by discussing how recent biological and technological advances should facilitate future studies into the functional roles of S-acylation–dependent signaling.An ever-growing number of studies highlight the importance of S-acylation, a reversible protein-lipid modification, for diverse aspects of intracellular signaling. In this review, we summarize the current understanding of how S-acylation regulates perhaps the best-known class of signaling enzymes, protein kinases. We describe how S-acylation acts as a membrane targeting signal that localizes certain kinases to specific membranes, and how such membrane localization in turn facilitates the assembly of signaling hubs consisting of an S-acylated kinase’s upstream activators and/or downstream targets. We further discuss recent findings that S-acylation can control additional aspects of the function of certain kinases, including their interactions and, surprisingly, their activity, and how such regulation might be exploited for potential therapeutic gain. We go on to describe the roles and regulation of de-S-acylases and how extracellular signals drive dynamic (de) S-acylation of certain kinases. We discuss how S-acylation has the potential to lead to “emergent properties” that alter the temporal profile and/or salience of intracellular signaling events. We close by giving examples of other S-acylation– dependent classes of signaling enzymes and by discussing how recent biological and technological advances should facilitate future studies into the functional roles of S-acylation–dependent signaling. [ABSTRACT FROM AUTHOR]

In type 1 diabetes (T1D), autoreactive immune cells infiltrate the pancreas and secrete proinflammatory cytokines that initiate cell death in insulin producing islet β-cells. Protein kinase C δ (PKCδ) plays a role in mediating cytokine-induced β-cell death; however, the exact mechanisms are not well understood. To address this, we used an inducible β-cell specific PKCδ KO mouse as well as a small peptide inhibitor of PKCδ. We identified a role for PKCδ in mediating cytokine-induced β-cell death and have shown that inhibiting PKCδ protects pancreatic β-cells from cytokine-induced apoptosis in both mouse and human islets. We determined that cytokines induced nuclear translocation and activity of PKCδ and that caspase-3 cleavage of PKCδ may be required for cytokine-mediated islet apoptosis. Further, cytokine activated PKCδ increases activity both of proapoptotic Bax with acute treatment and C-Jun N-terminal kinase with prolonged treatment. Overall, our results suggest that PKCδ mediates cytokine-induced apoptosis via nuclear translocation, cleavage by caspase-3, and upregulation of proapoptotic signaling in pancreatic β-cells. Combined with the protective effects of PKCδ inhibition with δV1-1, the results of this study will aid in the development of novel therapies to prevent or delay β-cell death and preserve β-cell function in T1D. [ABSTRACT FROM AUTHOR]

During cryopreservation, a substantial portion of spermatozoa undergoes apoptosis due to cryoinjury, resulting in decreased fertility. Boar spermatozoa are highly sensitive to temperature, with low temperature triggering reactive oxygen species (ROS) generation, leading to oxidative stress and apoptosis. Sulforaphane (SFN), a potent natural compound found in cruciferous vegetables, is efficacious in mitigating oxidative stress. We here supplemented different SFN concentrations (0, 1.25, 2.5, 5, 10, and 20 μM) into the freezing extender to explore its effect on boar sperm during cryopreservation and determine the optimal SFN concentration. Supplementation of 5 μM SFN exhibited the highest sperm motility, motion performance, plasma membrane integrity, acrosome integrity, and antioxidant properties (total antioxidant capacity (T-AOC) and antioxidant enzyme activity) after freezing and thawing. Then, RT group, C group and C + SFN group were established to explore the effect of SFN on the cryopreservation-induced sperm apoptosis level and fertilizing capacity of post-thawed sperms. SFN effectively rescued the apoptosis and fertilizing capacity of post-thawed sperms. Mechanistically, SFN activated the redox-sensitive nuclear factor erythroid 2-related factor 2 (NRF2/NFE2L2) by promoting adenosine monophosphate-activated protein kinase (AMPK) phosphorylation. This activation improved antioxidant defenses, ultimately improving cryoinjury in boar spermatozoa. In summary, SFN suppressed cryopreservation-induced apoptosis of spermatozoa by activating antioxidant defenses and the AMPK/NFE2L2 signaling pathway. These findings suggest a novel approach for augmenting the cryoprotective efficiency and spermatozoa fertility after cryopreservation. • SFN improved the quality of boar spermatozoa after cryopreservation. • SFN increased the antioxidant capacity of boar spermatozoa after cryopreservation. • SFN attenuated boar spermatozoa apoptosis induced by cryopreservation. • SFN exerts the beneficial effects by activating the NFE2L2/AMPK signaling pathway. [ABSTRACT FROM AUTHOR]

TOR protein kinases serve as the catalytic subunit of the TORC1 and TORC2 complexes, which regulate cellular growth, proliferation, and survival. In the fission yeast, Schizosaccharomyces pombe, cells lacking TORC2 or its downstream kinase Gad8 (AKT or SGK1 in human cells) exhibit sensitivity to a wide range of stress conditions, including DNA damage stress. One of the first responses to DNA damage is the phosphorylation of C-terminal serine residues within histone H2AX in human cells (γH2AX), or histone H2A in yeast cells (γH2A). The kinases responsible for γH2A in S. pombe are the two DNA damage checkpoint kinases Rad3 and Tel1 (ATR and ATM, respectively, in human cells). Here we report that TORC2-Gad8 signaling is required for accumulation of γH2A in response to DNA damage and during quiescence. Using the TOR-specific inhibitor, Torin1, we demonstrate that the effect of TORC2 on γH2A in response to DNA damage is immediate, rather than adaptive. The lack of γH2A is restored by deletion mutations of transcription and chromatin modification factors, including loss of components of Paf1C, SAGA, Mediator, and the bromo-domain proteins Bdf1/Bdf2. Thus, we suggest that TORC2-Gad8 may affect the accumulation of γH2A by regulating chromatin structure and function. [ABSTRACT FROM AUTHOR]

Isoforms of microtubule-associated protein 2 (MAP2) differ from their homolog Tau in the sequence and interactions of the N-terminal region. Binding of the N-terminal region of MAP2c (N-MAP2c) to the dimerization/docking domains of the regulatory subunit RIIa of cAMP-dependent protein kinase (RIIDD2) and to the Src-homology domain 2 (SH2) of growth factor receptor-bound protein 2 (Grb2) have been described long time ago. However, the structural features of the complexes remained unknown due to the disordered nature of MAP2. Here, we provide structural description of the complexes. We have solved solution structure of N-MAP2c in complex with RIIDD2, confirming formation of an amphiphilic α-helix of MAP2c upon binding, defining orientation of the ahelix in the complex and showing that its binding register differs from previous predictions. Using chemical shift mapping, we characterized the binding interface of SH2-Grb2 and rat MAP2c phosphorylated by the tyrosine kinase Fyn in their complex and proposed a model explaining differences between SH2-Grb2 complexes with rat MAP2c and phosphopeptides with a Grb2-specific sequence. The results provide the structural basis of a potential role of MAP2 in regulating cAMP-dependent phosphorylation cascade via interactions with RIIDD2 and Ras signaling pathway via interactions with SH2-Grb2. [ABSTRACT FROM AUTHOR]

In Saccharomyces cerevisiae, Pah1 phosphatidate (PA) phosphatase, which catalyzes the Mg2+-dependent dephosphorylation of PA to produce diacylglycerol, plays a key role in utilizing PA for the synthesis of the neutral lipid triacylglycerol and thereby controlling the PA-derived membrane phospholipids. The enzyme function is controlled by its subcellular location as regulated by phosphorylation and dephosphorylation. Pah1 is initially inactivated in the cytosol through phosphorylation by multiple protein kinases and then activated via its recruitment and dephosphorylation by the protein phosphatase Nem1-Spo7 at the nuclear/endoplasmic reticulum membrane where the PA phosphatase reaction occurs. Many of the protein kinases that phosphorylate Pah1 have yet to be characterized with the identification of the target residues. Here, we established Pah1 as a bona fide substrate of septinassociated Hsl1, a protein kinase involved in mitotic morphogenesis checkpoint signaling. The Hsl1 activity on Pah1 was dependent on reaction time and the amounts of protein kinase, Pah1, and ATP. The Hsl1 phosphorylation of Pah1 occurred on Ser-748 and Ser-773, and the phosphorylated protein exhibited a 5-fold reduction in PA phosphatase catalytic efficiency. Analysis of cells expressing the S748A and S773A mutant forms of Pah1 indicated that Hsl1-mediated phosphorylation of Pah1 promotes membrane phospholipid synthesis at the expense of triacylglycerol, and ensures the dependence of Pah1 function on the Nem1-Spo7 protein phosphatase. This work advances the understanding of how Hsl1 facilitates membrane phospholipid synthesis through the phosphorylation-mediated regulation of Pah1. [ABSTRACT FROM AUTHOR]

The PKC-related kinases (PRKs, also termed PKNs) are important in cell migration, cancer, hepatitis C infection, and nutrient sensing. They belong to a group of protein kinases called AGC kinases that share common features like a C-terminal extension to the catalytic domain comprising a hydrophobic motif. PRKs are regulated by N-terminal domains, a pseudosubstrate sequence, Rho-binding domains, and a C2 domain involved in inhibition and dimerization, while Rho and lipids are activators. We investigated the allosteric regulation of PRK2 and its interaction with its upstream kinase PDK1 using a chemical biology approach. We confirmed the phosphoinositide-dependent protein kinase 1 (PDK1)-interacting fragment (PIF)-mediated docking interaction of PRK2 with PDK1 and showed that this interaction can be modulated allosterically. We showed that the polypeptide PIFtide and a small compound binding to the PIF-pocket of PRK2 were allosteric activators, by displacing the pseudosubstrate PKL region from the active site. In addition, a small compound binding to the PIF-pocket allosterically inhibited the catalytic activity of PRK2. Together, we confirmed the docking interaction and allostery between PRK2 and PDK1 and described an allosteric communication between the PIF-pocket and the active site of PRK2, both modulating the conformation of the ATP-binding site and the pseudosubstrate PKL-binding site. Our study highlights the allosteric modulation of the activity and the conformation of PRK2 in addition to the existence of at least two different complexes between PRK2 and its upstream kinase PDK1. Finally, the study highlights the potential for developing allosteric drugs to modulate PRK2 kinase conformations and catalytic activity. [ABSTRACT FROM AUTHOR]

Protein phosphorylation by kinases regulates mammalian cell functions, such as growth, division, and signal transduction. Among human kinases, NME1 and NME2 are associated with metastatic tumor suppression but remain understudied due to the lack of tools to monitor their cellular substrates. In particular, NME1 and NME2 are multispecificity kinases phosphorylating serine, threonine, histidine, and aspartic acid residues of substrate proteins, and the heat and acid sensitivity of phosphohistidine and phosphoaspartate complicates substrate discovery and validation. To provide new substrate monitoring tools, we established the g-phosphate– modified ATP analog, ATP-biotin, as a cosubstrate for phosphorylbiotinylation of NME1 and NME2 cellular substrates. Building upon this ATP-biotin compatibility, the Kinasecatalyzed Biotinylation with Inactivated Lysates for Discovery of Substrates method enabled validation of a known substrate and the discovery of seven NME1 and three NME2 substrates. Given the paucity of methods to study kinase substrates, ATPbiotin and the Kinase-catalyzed Biotinylation with Inactivated Lysates for Discovery of Substrates method are valuable tools to characterize the roles of NME1 and NME2 in human cell biology. [ABSTRACT FROM AUTHOR]

KRAS-mutant cancers, due to their protein targeting complexity, present significant therapeutic hurdles. The identification of the macropinocytic phenotype in these cancers has emerged as a promising alternative therapeutic target. Our study introduces MPD1, an macropinocytosis-targeting peptide-drug conjugates (PDC), which is developed to treat KRAS mutant cancers. This PDC is specifically designed to trigger a positive feedback loop through its caspase-3 cleavable characteristic. However, we observe that this loop is hindered by DNA-PK mediated DNA damage repair processes in cancer cells. To counter this impediment, we employ AZD7648, a DNA-PK inhibitor. Interestingly, the combined treatment of MPD1 and AZD7648 resulted in a 100% complete response rate in KRAS-mutant xenograft model. We focus on the synergic mechanism of it. We discover that AZD7648 specifically enhances macropinocytosis in KRAS-mutant cancer cells. Further analysis uncovers a significant correlation between the increase in macropinocytosis and PI3K signaling, driven by AMPK pathways. Also, AZD7648 reinforces the positive feedback loop, leading to escalated apoptosis and enhanced payload accumulation within tumors. AZD7648 possesses broad applications in augmenting nano-sized drug delivery and preventing DNA repair resistance. The promising efficacy and evident synergy underscore the potential of combining MPD1 with AZD7648 as a strategy for treating KRAS-mutant cancers. [Display omitted] • Demonstrated enhancement of macropinocytosis mediated by DNA-PK inhibitor. • Uncovered the mechanism behind macropinocytosis induction via nutrition signaling. • Revealed PI3K mediated cancer-specific macropinocytosis increment. • Successfully operated an in situ tumor-eradicating positive feedback loop. • Achieved 100% CR of KRAS mutant tumors through systemic drug administration. [ABSTRACT FROM AUTHOR]

Reactive oxygen species (ROS) are considered toxic substances causing oxidative damage in plants, but they are also important signaling molecules crucial for many developmental processes of root and leaf. The origin of many major ROS signaling target proteins is highly conserved, predating the evolution of land plants. ROS function for the regulation of root and leaf development with the presence of phytohormones, gasotransmitters, and target proteins, particularly mitogen-activated protein kinases (MAPKs). Reactive oxygen species (ROS) are the key players in regulating developmental processes of plants. Plants have evolved a large array of gene families to facilitate the ROS-regulated developmental process in roots and leaves. However, the cellular targets of ROS during plant evolutionary development are still elusive. Here, we found early evolution and large expansions of protein families such as mitogen-activated protein kinases (MAPK) in the evolutionarily important plant lineages. We review the recent advances in interactions among ROS, phytohormones, gasotransmitters, and protein kinases. We propose that these signaling molecules act in concert to maintain cellular ROS homeostasis in developmental processes of root and leaf to ensure the fine-tuning of plant growth for better adaptation to the changing climate. [ABSTRACT FROM AUTHOR]

Synaptic dysfunction is highly correlated with cognitive impairments in Alzheimer's disease (AD), the most common dementia syndrome in the elderly. Long-term potentiation (LTP) and long-term depression (LTD) are two primary forms of synaptic plasticity with opposite direction of synaptic efficiency change. Both LTD and LTD are considered to mediate the cellular process of learning and memory. Substantial studies demonstrate AD-associated deficiency of both LTP and LTD. Meanwhile, the molecular signaling mechanisms underlying impairment of synaptic plasticity, particularly LTD, are poorly understood. By taking advantage of the novel transgenic mouse models recently developed in our lab, here we aimed to investigate the roles of AMP-activated protein kinase (AMPK), a central molecular senor that plays a critical role in maintaining cellular energy homeostasis, in regulation of LTD phenotypes in AD. We found that brain-specific suppression of the AMPKα1 isoform (but not AMPKα2 isoform) was able to alleviate mGluR-LTD deficits in APP/PS1 AD mouse model. Moreover, suppression of either AMPKα isoform failed to alleviate AD-related NMDAR-dependent LTD deficits. Taken together with our recent studies on roles of AMPK signaling in AD pathophysiology, the data indicate isoform-specific roles of AMPK in mediating AD-associated synaptic and cognitive impairments. • Hippocampal mGluR-LTD and NMDAR-LTD are impaired in aged APP/PS1 mice. • Genetic reduction of neuronal AMPKα1 alleviates mGluR-LTD failure in APP/PS1 mice. • Suppression of AMPKα isoforms does not alter NMDAR-LTD in APP/PS1 mice. [ABSTRACT FROM AUTHOR]

Protein tyrosine phosphorylation is a post-translational modification that regulates protein structure to modulate demic organisms' homeostasis and function. This physiological process is regulated by two enzyme families, protein tyrosine kinases (PTKs) and protein tyrosine phosphatases (PTPs). As an important regulator of protein function, PTPs are indispensable for maintaining cell intrinsic physiology in different systems, as well as liver physiological and pathological processes. Dysregulation of PTPs has been implicated in multiple liver-related diseases, including chronic liver diseases (CLDs), hepatocellular carcinoma (HCC), and liver injury, and several PTPs are being studied as drug therapeutic targets. Therefore, given the regulatory role of PTPs in diverse liver diseases, a collated review of their function and mechanism is necessary. Moreover, based on the current research status of targeted therapy, we emphasize the inclusion of several PTP members that are clinically significant in the development and progression of liver diseases. As an emerging breakthrough direction in the treatment of liver diseases, this review summarizes the research status of PTP-targeting compounds in liver diseases to illustrate their potential in clinical treatment. Overall, this review aims to support the development of novel PTP-based treatment pathways for liver diseases. Protein tyrosine phosphatases are involved in liver disease progression by catalyzing substrates dephosphorylation and some of them are potential therapeutic targets. [Display omitted] [ABSTRACT FROM AUTHOR]

• Guava leaf extract (GLE) enhanced muscle strength in muscle atrophy (MA) mice. • GLE normalized abnormal morphological changes in MA mice. • GLE attenuated muscle proteolysis via ubiquitin proteasome system in MA mice. • GLE reduced excessive autophagy and apoptosis, which triggers skeletal muscle damage in MA mice. Muscle atrophy is the waste or loss of muscle mass and is caused by physical inactivity, aging, or diseases such as diabetes, cancer, and heart failure. The number of patients suffering from musculoskeletal disorders is expected to increase in the future. However, intervention for muscle atrophy is limited, so research on treatment for muscle wasting is needed. This study hypothesized that guava leaf (Psidium guajava L. [GL]) would have ameliorative effects on muscle atrophy by regulation of protein degradation pathways in a dexamethasone (DEX)-induced muscle atrophy mice model. Muscle atrophy was induced by DEX injection for 28 days in 7 week-old-male ICR mice. Then, low-dose GL (LGL, 200 mg/kg) or high-dose GL (HGL, 500 mg/kg) extract (GLE) was supplemented by oral gavage for 21 days. Muscle strength, calf thickness, and body composition were analyzed. Histopathological changes in the gastrocnemius muscle were examined using hematoxylin and eosin staining, and molecular pathways related to muscle degradation were analyzed by western blots. GLE treatment regardless of dose increased muscle strength in mice with muscle atrophy accompanied by attenuating autophagy related pathway in the DEX-induced muscle atrophy mice. Moreover, a high dose of GLE treatment ameliorated ubiquitin proteasome system and apoptosis in the DEX-induced muscle atrophy mice. This study suggested that GLE could be helpful to improve muscle health and alleviate proteolysis by regulation of the ubiquitin–proteasome system, autophagy, and apoptosis, which are involved in muscle degradation. In conclusion, GLE could be a potential nutraceutical to prevent muscle atrophy. GLE contains many types of polyphenols such as gallic acid, epicatechin, quercetin, caffeic acid, and others. GLE supplementation attenuated muscle loss and enhanced muscle strength by inhibition of UPS-mediated muscle degradation, mTOR-related autophagy, and apoptosis pathway in DEX-induced muscle atrophy mice. Consequently, GLE might attenuate muscle dysfunction through multiple mechanisms expected to be used to prevent muscle atrophy. Abbreviations: CSA, cross-sectional area; DEX, dexamethasone; GL, guava leaf; GLE, GL extract; UPS, ubiquitin–proteasome system [Display omitted] [ABSTRACT FROM AUTHOR]

Traumatic brain injury (TBI) is an important health concern in the society. Previous studies have suggested that necroptosis occurs following TBI. However, the underlying mechanisms and roles of necroptosis are not well understood. In this study, we aimed to assess the role of receptor-interacting serine/threonine-protein kinase 3 (RIP3)-mediated necroptosis after TBI both in vitro and in vivo. We established a cell-stretching injury and mouse TBI model by applying a cell injury controller and controlled cortical impactor to evaluate the relationships among necroptosis, apotosis, inflammation, and TBI both in vitro and in vivo. The results revealed that necroptosis mediated by RIP1, RIP3, and mixed lineage kinase domain-like protein was involved in secondary TBI. Additionally, protein kinase B (Akt), phosphorylated Akt, mammalian target of rapamycin (mTOR), and phosphorylated mTOR potentially contribute to necroptosis. The inhibition of RIP3 by GSK'872 (a specific inhibitor) blocked necroptosis and reduced the activity of Akt/mTOR, leading to the alleviation of inflammation by reducing the levels of NOD-, LRR- and pyrin domain-containing protein 3. Moreover, the inhibition of RIP3 by GSK'872 promoted the activity of cysteinyl aspartate specific proteinase-8, an enzyme involved in apoptosis and inflammation. These data demonstrate that RIP3 inhibition could improve the prognosis of TBI, based on the attenuation of inflammation by switching RIP3-dependent necroptosis to cysteinyl aspartate specific proteinase-8-dependent apoptosis. [ABSTRACT FROM AUTHOR]

[Display omitted] • The iGG analysis has been proposed to utilize evolutionary information between the crop and several its relatives. • The evolution of CIPK and CBL genes have been characterized in seven species to prove the iGG method. • The divergent expression is important for keeping the stoichiometric balance of TaCBL-TaCIPK. • Our iGG analysis reveals a Triticeae-specific TaCIPK duplicate responding to drought. • Transgenic plants of TaCIPK17 -OE in tobacco and wheat showed higher resistance to drought. Reverse genetic studies conducted in the plant with a complex or polyploidy genome enriched with large gene families (like wheat) often meet challenges in identifying the key candidate genes related to important traits and prioritizing the genes for functional experiments. To overcome the above-mentioned challenges of reverse genetics, this work aims to establish an efficient multi-species strategy for genome-wide gene identification and prioritization of the key candidate genes. We established the integrative gene duplication and genome-wide analysis (iGG analysis) as a strategy for pinpointing key candidate genes deserving functional research. The iGG captures the evolution, and the expansion/contraction of large gene families across phylogeny-related species and integrates spatial–temporal expression information for gene function inference. Transgenic approaches were also employed to functional validation. As a proof-of-concept for the iGG analysis, we took the wheat calcineurin B-like protein-interacting protein kinases (CIPKs) family as an example. We identified CIPK s from seven monocot species, established the orthologous relationship of CIPK s between rice and wheat, and characterized Triticeae-specific CIPK duplicates (e.g. , CIPK4 and CIPK17). Integrated with our analysis of CBLs and CBL-CIPK interaction, we revealed that divergent expressions of TaCBL s and TaCIPK s could play an important role in keeping the stoichiometric balance of CBL-CIPK. Furthermore, we validated the function of TaCIPK17-A2 in the regulation of drought tolerance by using transgenic approaches. Overexpression of TaCIPK17 enhanced antioxidant capacity and improved drought tolerance in wheat. The iGG analysis leverages evolutionary and comparative genomics of crops with large genomes to rapidly highlight the duplicated genes potentially associated with speciation, domestication and/or particular traits that deserve reverse-genetic functional studies. Through the identification of Triticeae-specific TaCIPK17 duplicates and functional validation, we demonstrated the effectiveness of the iGG analysis and provided a new target gene for improving drought tolerance in wheat. [ABSTRACT FROM AUTHOR]

Recently, significant advances have been achieved in gene editing of plant protein kinases and phosphatases using clustered regularly interspaced short palindromic repeats/CRISPR-associated protein 9 (CRISPR/Cas9) technology. Progress in computer science and programming could significantly improve CRISPR/Cas9 gene-editing predictions and subsequent wet-laboratory experiments on protein kinases and phosphatases. Gene-edited protein kinases and phosphatases could considerably improve desirable crop traits and stress tolerance. Protein phosphorylation, the most common and essential post-translational modification, belongs to crucial regulatory mechanisms in plants, affecting their metabolism, intracellular transport, cytoarchitecture, cell division, growth, development, and interactions with the environment. Protein kinases and phosphatases, two important families of enzymes optimally regulating phosphorylation, have now become important targets for gene editing in crops. We review progress on gene-edited protein kinases and phosphatases in crops using clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR-associated protein 9 (Cas9). We also provide guidance for computational prediction of alterations and/or changes in function, activity, and binding of protein kinases and phosphatases as consequences of CRISPR/Cas9-based gene editing with its possible application in modern crop molecular breeding towards sustainable agriculture. [ABSTRACT FROM AUTHOR]

Type I interferon (IFN) inhibits a wide spectrum of viruses through stimulating the expression of antiviral proteins. As an IFN-induced protein, myxovirus resistance B (MXB) protein was reported to inhibit multiple highly pathogenic human viruses. It remains to be determined whether MXB employs a common mechanism to restrict different viruses. Here, we find that IFN alters the subcellular localization of hundreds of host proteins, and this IFN effect is partially lost upon MXB depletion. The results of our mechanistic study reveal that MXB recognizes vimentin (VIM) and recruits protein kinase B (AKT) to phosphorylate VIM at amino acid S38, which leads to reorganization of the VIM network and impairment of intracellular trafficking of virus protein complexes, hence causing a restriction of virus infection. These results highlight a new function of MXB in modulating VIM-mediated trafficking, which may lead towards a novel broad-spectrum antiviral strategy to control a large group of viruses that depend on VIM for successful replication. MXB recruits AKT to phosphorylate vimentin at S38, which leads to the rearrangement of vimentin and impairment of intracellular trafficking of virus protein complexes. [Display omitted] [ABSTRACT FROM AUTHOR]

In neurodegeneration, neurons release lipids that accumulate in glial lipid droplets (LDs). But what controls lipid transport and how does this affect glia? A recent study by Li et al. discovered that the loss of neuronal AMP-activated protein kinase (AMPK) activity promotes lipid efflux, which drives a proinflammatory state in microglia. [ABSTRACT FROM AUTHOR]

Since chlormequat chloride is widely applied as a plant growth regulator in agriculture and horticulture, its exposure through food consumption is common. We demonstrated previously that chlormequat chloride exposure during pregnancy led to embryos with bigger sizes associated with higher levels of growth hormone (GH) on gestation day 11 (GD11). However, the dose-effect relationship of chlormequat chloride at a lower dose range was not established, and the underlying mechanisms of its promoting effects on embryonic growth and development were not fully elucidated. To address these, pregnant rats were orally exposed to chlormequat chloride at 0, 0.05, 0.5 and 5 mg/kg.bw from GD0 to 11 and the embryonic growth and growth related hormones were evaluated on GD11. We found that the growth and development of the embryos was significantly promoted in a dose dependent manner by chlormequat chloride. Chlormequat chloride also increased embryonic GH, GH releasing hormone (GHRH), and somatostatin (SRIF), and inhibited the embryonic cAMP dependent protein kinase A (PKA) signaling pathway. Chlormequat chloride increased GH synthesis modulated by GHRH/SRIF-PKA-Pituitary specific transcription factor 1 (Pit-1) in the maternal rats. Intriguingly, chlormequat chloride did not show any effects on GH and PKA signaling pathways in the non-pregnant female rats. These findings together suggest that the disrupting effect of chlormequat chloride on GH is associated with pregnancy. • Chlormequat exposure at ADI during pregnancy stimulated the growth and development of the embryos on GD11. • Chlormequat increased embryonic GH, GHRH, and SRIF, while inhibited the PKA signaling pathway in the embryos. • Chlormequat treatment led to increased GH synthesis mediated by GHRH/SRIF-PKA-Pit-1 in the maternal pituitary. • chlormequat chloride did not affect GH and its related PKA signaling pathways in the non-pregnant female rats. [ABSTRACT FROM AUTHOR]

Methicillin-resistant Staphylococcus aureus (MRSA) is the primary pathogenic agent responsible for epidermal wound infection and suppuration, seriously threatening the life and health of human beings. To address this fundamental challenge, we propose a heterojunction nanocomposite (Ca-CN/MnS) comprised of Ca-doped g-C 3 N 4 and MnS for the therapy of MRSA-accompanied wounds. The Ca doping leads to a reduction in both the bandgap and the singlet state S 1 –triplet state T 2 energy gap (Δ E ST). The Ca doping also facilitates the two-photon excitation, thus remarkably promoting the separation and transfer of 808 nm near-infrared (NIR) light–triggered electron–hole pairs together with the built-in electric field. Thereby, the production of reactive oxygen species and heat are substantially augmented nearby the nanocomposite under 808 nm NIR light irradiation. Consequently, an impressive photocatalytic MRSA bactericidal efficiency of 99.98 ± 0.02 % is achieved following exposure to NIR light for 20 min. The introduction of biologically functional elements (Ca and Mn) can up-regulate proteins such as pyruvate kinase (PKM), L-lactate dehydrogenase (LDHA), and calcium/calmodulin-dependent protein kinase (CAMKII), trigger the glycolysis and calcium signaling pathway, promote cell proliferation, cellular metabolism, and angiogenesis, thereby expediting the wound-healing process. This heterojunction nanocomposite, with its precise charge-transfer pathway, represents a highly effective bactericidal and bioactive system for treating multidrug-resistant bacterial infections and accelerating tissue repair. Due to the bacterial resistance, developing an antibiotic-free and highly effective bactericidal strategy to treat bacteria-infected wounds is critical. We have designed a heterojunction consisting of calcium doped g-C 3 N 4 and MnS (Ca-CN/MnS) that can rapidly kill methicillin-resistant Staphylococcus aureus (MRSA) without damaging normal tissue through a synergistic effect of two-photon stimulated photothermal and photodynamic therapy. In addition, the release of trace amounts of biofunctional elements Mn and Ca triggers glycolysis and calcium signaling pathways that promote cellular metabolism and cell proliferation, contributing to tissue repair and wound healing. [Display omitted] [ABSTRACT FROM AUTHOR]

Chemoresistance remains a major challenge in treating pancreatic ductal adenocarcinoma (PDAC). Although chemoradiation has proven effective in other tumor types, such as head and neck squamous cell carcinoma, its role in PDAC and effect on acquired chemoresistance have yet to be fully explored. In this study, we investigated the sensitivity of gemcitabine-resistant (GR) and paclitaxel-resistant (PR) PDAC cells to ionizing radiation (IR) and their underlying mechanisms. GR and PR clones were generated from PANC-1, PATU-T, and SUIT2-007 pancreatic cancer cell lines. Cell survival after radiation was assessed using clonogenic assay, sulforhodamine B assay, apoptosis, and spheroid growth by bioluminescence. Radiation-induced DNA damage was assessed using Western blot, extra-long polymerase chain reaction, reactive oxygen species production, and immunofluorescence. Autophagy and modulation of the Hippo signaling pathway were investigated using proteomics, Western blot, immunofluorescence, and reverse-transcription quantitative polymerase chain reaction. In both 2- and 3-dimensional settings, PR cells were more sensitive to IR and showed decreased β-globin amplification, indicating more DNA damage accumulation compared with GR or wild-type cells after 24 hours. Proteomic analysis of PR PATU-T cells revealed that the protein MST4, a kinase involved in autophagy and the Hippo signaling pathway, was highly downregulated. A differential association was found between autophagy and radiation treatment depending on the cell model. Interestingly, increased yes-associated protein nuclear localization and downstream Hippo signaling pathway target gene expression were observed in response to IR. This was the first study investigating the potential of IR in targeting PDAC cells with acquired chemoresistance. Our results demonstrate that PR cells exhibit enhanced sensitivity to IR due to greater accumulation of DNA damage. Additionally, depending on the specific cellular context, radiation-induced modulation of autophagy and the Hippo signaling pathway emerged as potential underlying mechanisms, findings with potential to inform personalized treatment strategies for patients with acquired chemoresistance. [ABSTRACT FROM AUTHOR]

Cell signaling occurs in the restricted confines of highly organized intracellular signaling nanodomains. Receptor-associated independent cAMP nanodomains (RAINs) drive local cAMP synthesis and localized signaling. Optical mapping of local cAMP pools has defined local cAMP nanodomains due to buffered diffusion of second messengers. G protein-coupled receptor (GPCR) signaling at subcellular membrane compartments generates isolated intracellular pools of cAMP that drive unique cellular functions. A-kinase anchoring protein (AKAP)-signaling islands regionally distributed throughout the cell potentiate individual protein kinase A (PKA) phosphorylation events at the nanometer scale. Louis Pasteur once famously said 'in the fields of observation chance favors only the prepared mind'. Much of chance is being in the right place at the right time. This is particularly true in the crowded molecular environment of the cell where being in the right place is often more important than timing. Although Brownian motion argues that enzymes will eventually bump into substrates, this probability is greatly enhanced if both molecules reside in the same subcellular compartment. However, activation of cell signaling enzymes often requires the transmission of chemical signals from extracellular stimuli to intracellular sites of action. This review highlights new developments in our understanding of cAMP generation and the 3D utilization of this second messenger inside cells. [ABSTRACT FROM AUTHOR]

Understanding the mechanisms that govern the stability of functionally crucial proteins is essential for various cellular processes, development, and overall cell viability. Disturbances in protein homeostasis are linked to the pathogenesis of neurodegenerative diseases. PTEN-induced kinase 1 (PINK1), a protein kinase, plays a significant role in mitochondrial quality control and cellular stress response, and its mutated forms lead to early-onset Parkinson's disease. Despite its importance, the specific mechanisms regulating PINK1 protein stability have remained unclear. This study reveals a cytoplasmic interaction between PINK1 and F-box and WD repeat domain--containing 7β (FBW7β) in mammalian cells. FBW7β, a component of the Skp1-Cullin-1-F-box protein complex--type ubiquitin ligase, is instrumental in recognizing substrates. Our findings demonstrate that FBW7β regulates PINK1 stability through the Skp1- Cullin-1-F-box protein complex and the proteasome pathway. It facilitates the K48-linked polyubiquitination of PINK1, marking it for degradation. When FBW7 is absent, PINK1 accumulates, leading to heightened mitophagy triggered by carbonyl cyanide 3-chlorophenylhydrazone treatment. Moreover, exposure to the toxic compound staurosporine accelerates PINK1 degradation via FBW7β, correlating with increased cell death. This study unravels the intricate mechanisms controlling PINK1 protein stability and sheds light on the novel role of FBW7β. These findings deepen our understanding of PINK1-related pathologies and potentially pave the way for therapeutic interventions. [ABSTRACT FROM AUTHOR]

Protein tyrosine kinases (RTKs) modulate a wide range of pathophysiological events in several non-malignant disorders, including diabetic complications. To find new targets driving the development of diabetic cardiomyopathy (DCM), we profiled an RTKs phosphorylation array in diabetic mouse hearts and identified increased phosphorylated fibroblast growth factor receptor 1 (p-FGFR1) levels in cardiomyocytes, indicating that FGFR1 may contribute to the pathogenesis of DCM. Using primary cardiomyocytes and H9C2 cell lines, we discovered that high-concentration glucose (HG) transactivates FGFR1 kinase domain through toll-like receptor 4 (TLR4) and c-Src, independent of FGF ligands. Knocking down the levels of either TLR4 or c-Src prevents HG-activated FGFR1 in cardiomyocytes. RNA-sequencing analysis indicates that the elevated FGFR1 activity induces pro-inflammatory responses via MAPKs–NF κ B signaling pathway in HG-challenged cardiomyocytes, which further results in fibrosis and hypertrophy. We then generated cardiomyocyte-specific FGFR1 knockout mice and showed that a lack of FGFR1 in cardiomyocytes prevents diabetes-induced cardiac inflammation and preserves cardiac function in mice. Pharmacological inhibition of FGFR1 by a selective inhibitor, AZD4547, also prevents cardiac inflammation, fibrosis, and dysfunction in both type 1 and type 2 diabetic mice. These studies have identified FGFR1 as a new player in driving DCM and support further testing of FGFR1 inhibitors for possible cardioprotective benefits. Hyperglycemia activates FGFR1 through TLR4 and c-Src axis in cardiomyopathy, which induces MAPKs-mediated NF κ B activation to increase inflammatory response and subsequent fibrosis and hypertrophy, driving the pathogenesis of DCM. [Display omitted] [ABSTRACT FROM AUTHOR]

Immune-mediated liver injury (ILI) is a condition where an aberrant immune response due to various triggers causes the destruction of hepatocytes. Fibroblast growth factor 4 (FGF4) was recently identified as a hepatoprotective cytokine; however, its role in ILI remains unclear. In patients with autoimmune hepatitis (type of ILI) and mouse models of concanavalin A (ConA)- or S-100-induced ILI, we observed a biphasic pattern in hepatic FGF4 expression, characterized by an initial increase followed by a return to basal levels. Hepatic FGF4 deficiency activated the mitochondria-associated intrinsic apoptotic pathway, aggravating hepatocellular apoptosis. This led to intrahepatic immune hyper-reactivity, inflammation accentuation, and subsequent liver injury in both ILI models. Conversely, administration of recombinant FGF4 reduced hepatocellular apoptosis and rectified immune imbalance, thereby mitigating liver damage. The beneficial effects of FGF4 were mediated by hepatocellular FGF receptor 4, which activated the Ca2+/calmodulin-dependent protein kinasekinase 2 (CaMKK β) and its downstream phosphatase and tensin homologue-induced putative kinase 1 (PINK1)-dependent B-cell lymphoma 2-like protein 1-isoform L (Bcl-X L) signalling axis in the mitochondria. Hence, FGF4 serves as an early response factor and plays a protective role against ILI, suggesting a therapeutic potential of FGF4 and its analogue for treating clinical immune disorder-related liver injuries. FGF4 is a responsive factor and potential therapeutic agent for immune liver injury (ILI). [Display omitted] [ABSTRACT FROM AUTHOR]

Bacterial infection was the major etiology for pulpal/root canal infection. This study aimed to investigate the activation of toll-like receptor-3 (TLR) on cyclooxygenase-2 (COX-2) expression and prostaglandin E 2 (PGE 2) and PGF 2α production of human dental pulp cells (HDPCs) and associated signaling. HDPCs were exposed to different concentrations of Poly (I:C) (a TLR3 activator). Cell viability was determined by 3- (4,5-Dimethylthiazol-2-yl)-2,5- diphenyltetrazolium bromide (MTT) assay and alkaline phosphatase (ALP) activity was evaluated by ALP staining. Activation of extracellular signal-regulated kinase (ERK) and p38 by Poly (I:C) was determined by immunofluorescent staining. The COX-2 protein expression was analyzed by Western blot. PGE 2 and PGF 2α production was measured by enzyme-linked immunosorbent assay. The mRNA expression was studied by real-time polymerase-chain reaction. Moreover, HDPCs were exposed to Poly(I:C) with/without U0126 or SB203580 treatment and analysis of COX-2 expression and prostanoid production were conducted. Poly (I:C) showed little effect on ALP activity, but decreased viability of HDPCs. It stimulated COX-2 mRNA and protein expression. Poly (I:C) induced PGE 2 and PGF 2α production of HDPCs. Poly (I:C) activated p -ERK, and p-p38 protein expression. Treatment by U0126 (a mitogen-activated protein kinase kinase (MEK)/ERK inhibitor) and SB203580 (a p38 inhibitor) attenuated Poly (I:C)-induced COX-2 mRNA and protein expression as well as PGE 2 and PGF 2α production. TLR3 activation is involved in the infection and inflammatory responses of pulp tissues, via MEK/ERK , and p38 signaling to mediate COX-2 expression as well as PGE 2 and PGF 2α production, contributing to the pathogenesis and progression of pulpal/periapical diseases. [ABSTRACT FROM AUTHOR]

The PAS (Per-ARNT-Sim) domain is a sensory protein regulatory module found in archaea, prokaryotes, and eukaryotes. Histidine and serine/threonine protein kinases, chemo- and photoreceptors, circadian rhythm regulators, ion channels, phosphodiesterases, and other cellular response regulators are among these proteins. Hik33 is a multifunctional sensory histidine kinase that is implicated in cyanobacterial responses to cold, salt, hyperosmotic, and oxidative stressors. The functional roles of individual Hik33 domains in signal transduction were investigated in this study. Synechocystis Hik33 deletion variants were developed, in which either both or a portion of the transmembrane domains and/or the PAS domain were deleted. Cold stress was applied to the mutant strains either under illumination or in the dark. The findings show that the transmembrane domains govern temperature responses, whereas PAS domain may be involved in regulation of downstream gene expression in light-dependent manner. [Display omitted] • The role of individual domains of Hik33 in light-dependent cold sensing have been studied. • PAS domain of Hik33 participates in light sensing. • Transmembrane domains of Hik33 are responsible for the perception of cold via sensing of changes in membrane thickness. [ABSTRACT FROM AUTHOR]

DNA-dependent protein kinase (DNA-PK) plays a key role in the repair of DNA double strand breaks via nonhomologous end joining. Inhibition of DNA-PK can enhance the effect of DNA double strand break inducing anticancer therapies. Peposertib (formerly "M3814") is an orally administered, potent, and selective small molecule DNA-PK inhibitor that has demonstrated radiosensitizing and antitumor activity in xenograft models and was well-tolerated in monotherapy. This phase 1 trial (National Clinical Trial 02516813) investigated the maximum tolerated dose, recommended phase 2 dose (RP2D), safety, and tolerability of peposertib in combination with palliative radiation therapy (RT) in patients with thoracic or head and neck tumors (arm A) and of peposertib in combination with cisplatin and curative-intent RT in patients with squamous cell carcinoma of the head and neck (arm B). Patients received peposertib once daily in ascending dose cohorts as a tablet or capsule in combination with palliative RT (arm A) or in combination with intensity modulated curative-intent RT and cisplatin (arm B). The most frequently observed treatment-emergent adverse events were radiation skin injury, fatigue, and nausea in arm A (n = 34) and stomatitis, nausea, radiation skin injury, and dysgeusia in arm B (n = 11). Based on evaluations of dose-limiting toxicities, tolerability, and pharmacokinetic data, RP2D for arm A was declared as 200 mg peposertib tablet once daily in combination with RT. In arm B (n = 11), 50 mg peposertib was declared tolerable in combination with curative-intent RT and cisplatin. However, enrollment was discontinued because of insufficient exposure at that dose, and the RP2D was not formally declared. Peposertib in combination with palliative RT was well-tolerated up to doses of 200 mg once daily as tablet with each RT fraction. When combined with RT and cisplatin, a tolerable peposertib dose yielded insufficient exposure. [ABSTRACT FROM AUTHOR]

Approximately 5 to 15% of nonmedullary thyroid cancers (NMTC) present in a familial form (familial nonmedullary thyroid cancers [FNMTC]). The genetic basis of FNMTC remains largely unknown, representing a limitation for diagnostic and clinical management. Recently, germline mutations in DNA repair-related genes have been described in cases with thyroid cancer (TC), suggesting a role in FNMTC etiology. Here, two FNMTC families were studied, each with two members affected with TC. Ninety-four hereditary cancer predisposition genes were analyzed through next-generation sequencing, revealing two germline CHEK2 missense variants (c.962A > C, p.E321A and c.470T > C, p.I157T), which segregated with TC in each FNMTC family. p.E321A, located in the CHK2 protein kinase domain, is a rare variant, previously unreported in the literature. Conversely, p.I157T, located in CHK2 forkhead-associated domain, has been extensively described, having conflicting interpretations of pathogenicity. CHK2 proteins (WT and variants) were characterized using biophysical methods, molecular dynamics simulations, and immunohistochemistry. Overall, biophysical characterization of theseCHK2variants showed that they have compromised structural and conformational stability and impaired kinase activity, compared to the WT protein. CHK2 appears to aggregate into amyloid-like fibrils in vitro, which opens future perspectives toward positioning CHK2 in cancer pathophysiology. CHK2 variants exhibited higher propensity for this conformational change, also displaying higher expression in thyroid tumors. The present findings support the utility of complementary biophysical and in silico approaches toward understanding the impact of genetic variants in protein structure and function, improving the current knowledge on CHEK2 variants' role in FNMTC genetic basis, with prospective clinical translation. [ABSTRACT FROM AUTHOR]

Resistance (R) genes in the Triticeae tribe include not only genes encoding the canonical intracellular nucleotide-binding leucine-rich-repeat proteins (NLRs) but also genes encoding kinase fusion proteins (KFPs). Exploring these unconventional KFPs may expand the scope of effector-triggered immunity (ETI) and will have significant implications for crop improvement. [ABSTRACT FROM AUTHOR]

In a recent study, Ziak et al. employed precise sparse labeling and spatiotemporally controlled genetic manipulations to uncover novel regulators of axon branching of layer 2/3 mouse callosal projection neurons. The authors elucidated a cell-autonomous signaling pathway wherein glycogen synthase kinase 3β (GSK3β) phosphorylation of microtubule-associated protein 1B (MAP1B) restricts interstitial axon branching by modulating microtubule (MT) tyrosination status. [ABSTRACT FROM AUTHOR]

• Neuraxial morphine increases spinal caspase-6 cleavage and PKMζ phosphorylation. • Caspase-6 inhibition reduces morphine-induced itch and PKMζ phosphorylation. • PKMζ inhibition attenuates morphine-induced acute itch. • Caspase-6 inhibition reduces chronic itch after contact dermatitis. Patients receiving neuraxial treatment with morphine for pain relief often experience a distressing pruritus. Neuroinflammation-mediated plasticity of sensory synapses in the spinal cord is critical for the development of pain and itch. Caspase-6, as an intracellular cysteine protease, is capable of inducing central nociceptive sensitization through regulating synaptic transmission and plasticity. Given the tight interaction between protein kinase Mζ (PKMζ) and excitatory synaptic plasticity, this pre-clinical study investigates whether caspase-6 contributes to morphine-induced itch and chronic itch via PKMζ. Intrathecal morphine and contact dermatitis were used to cause pruritus in mice. Morphine antinociception, itch-induced scratching behaviors, spinal activity of caspase-6, and phosphorylation of PKMζ and ERK were examined. Caspase-6 inhibitor Z-VEID-FMK, exogenous caspase-6 and PKMζ inhibitor ZIP were utilized to reveal the mechanisms and prevention of itch. Herein, we report that morphine induces significant scratching behaviors, which is accompanied by an increase in spinal caspase-6 cleavage and PKMζ phosphorylation (but not expression). Intrathecal injection of Z-VEID-FMK drastically reduces morphine-induced scratch bouts and spinal phosphorylation of PKMζ, without abolishing morphine analgesia. Moreover, intrathecal strategies of ZIP dose-dependently reduce morphine-induced itch-like behaviors. Spinal phosphorylation of ERK following neuraxial morphine is down-regulated by ZIP therapy. Recombinant caspase-6 directly exhibits scratching behaviors and spinal phosphorylation of ERK, which is compensated by PKMζ inhibition. Also, spinal inhibition of caspase-6 and PKMζ reduces the generation and maintenance of dermatitis-induced chronic itch. Together, these findings demonstrate that spinal caspase-6 modulation of PKMζ phosphorylation is important in the development of morphine-induced itch and dermatitis-induced itch in mice. [ABSTRACT FROM AUTHOR]

Major depressive disorder (MDD) remains a significant global health concern, with limited and slow efficacy of existing antidepressants. Electroconvulsive therapy (ECT) has superior and immediate efficacy for MDD, but its action mechanism remains elusive. Therefore, the elucidation of the action mechanism of ECT is expected to lead to the development of novel antidepressants with superior and immediate efficacy. Recent studies suggest a potential role of hippocampal astrocyte in MDD and ECT. Hence, we investigated antidepressant effect of electroconvulsive stimulation (ECS), an animal model of ECT, -related genes in hippocampal astrocyte with a mouse model of MDD, in which corticosterone (CORT)-induced depression-like behaviors were recovered by ECS. In this model, both of CORT-induced depression-like behaviors and the reduction of hippocampal astrocyte were recovered by ECS. Following it, astrocytes were isolated from the hippocampus of this model and RNA-seq was performed with these isolated astrocytes. Interestingly, gene expression patterns altered by CORT were reversed by ECS. Additionally, cell proliferation-related signaling pathways were inhibited by CORT and recovered by ECS. Finally, serum and glucocorticoid kinase-1 (SGK1), a multi-functional protein kinase, was identified as a candidate gene reciprocally regulated by CORT and ECS in hippocampal astrocyte. Our findings suggest a potential role of SGK1 in the antidepressant effect of ECS via the regulation of the proliferation of astrocyte and provide new insights into the involvement of hippocampal astrocyte in MDD and ECT. Targeting SGK1 may offer a novel approach to the development of new antidepressants which can replicate superior and immediate efficacy of ECT. [ABSTRACT FROM AUTHOR]

Evidence has suggested that addiction and memory systems are related, but the signaling cascades underlying this interaction have not been completelyealed yet. The importance of calcium-calmodulin-dependent protein kinase II (CaMKII) and brain-derived neurotrophic factor (BDNF) in the memory processes and also in drug addiction has been previously established. In this present investigation, we examined the effects of repeated morphine pretreatment on impairment of spatial learning and memory acquisition induced by systemic ethanol administration in adult male rats. Also, we assessed how these drug exposures influence the expression level of CaMKII and BDNF in the hippocampus and amygdala. Animals were trained by a single training session of 8 trials, and a probe test containing a 60-s free-swim without a platform was administered 24 h later. Before training trials, rats were treated with a once-daily subcutaneous morphine injection for 3 days followed by a 5-day washout period. The results showed that pre-training ethanol (1 g/kg) impaired spatial learning and memory acquisition and down-regulated the mRNA expression of CaMKII and BDNF. The amnesic effect of ethanol was suppressed in morphine- (15 mg/kg/day) pretreated animals. Furthermore, the mRNA expression level of CaMKII and BDNF increased significantly following ethanol administration in morphine-pretreated rats. Conversely, this improvement in spatial memory acquisition was prevented by daily subcutaneous administration of naloxone (2 mg/kg) 15 min prior to morphine administration. Our findings suggest that sub-chronic morphine treatment reverses ethanol-induced spatial memory impairment, which could be explained by modulating CaMKII and BDNF mRNA expressions in the hippocampus and amygdala. [Display omitted] • Pre-training acute administration of ethanol impaired spatial memory. • Amnesia induced by ethanol was decreased by morphine pre-exposure. • Ethanol increased CaMKII in the hippocampus and amygdala in morphine-pretreated rats. • Ethanol increased BDNF in the hippocampus and amygdala in morphine-pretreated rats. [ABSTRACT FROM AUTHOR]

The yeast Snf1/AMP-activated kinase (AMPK) maintains energy homeostasis, controlling metabolic processes and glucose derepression in response to nutrient levels and environmental cues. Under conditions of nitrogen or glucose limitation, Snf1 regulates pseudohyphal growth, a morphological transition characterized by the formation of extended multi-cellular filaments. During pseudohyphal growth, Snf1 is required for wild-type levels of inositol polyphosphate (InsP), soluble phosphorylated species of the six-carbon cyclitol inositol that function as conserved metabolic second messengers. InsP levels are established through the activity of a family of inositol kinases, including the yeast inositol polyphosphate kinase Kcs1, which principally generates pyrophosphorylated InsP7. Here, we report that Snf1 regulates Kcs1, affecting Kcs1 phosphorylation and inositol kinase activity. A snf1 kinase-defective mutant exhibits decreased Kcs1 phosphorylation, and Kcs1 is phosphorylated in vivo at Ser residues 537 and 646 during pseudohyphal growth. By in vitro analysis, Snf1 directly phosphorylates Kcs1, predominantly at amino acids 537 and 646. A yeast strain carrying kcsl encoding Ser-to-Ala point mutations at these residues (kcs1-S537A,S646A) shows elevated levels of pyrophosphorylated InsP7, comparable to InsP7 levels observed upon deletion of SNF1. The kcsl -S537A,S646A mutant exhibits decreased pseudohyphal growth, invasive growth, and cell elongation. Transcriptional profiling indicates extensive perturbation of metabolic pathways in kcs1-S537A,S646A. Growth of kcs1-S537A,S646A is affected on medium containing sucrose and antimycin A, consistent with decreased Snf1p signaling. This work identifies Snf1 phosphorylation of Kcs1, collectively highlighting the interconnectedness of AMPK activity and InsP signaling in coordinating nutrient availability, energy homoeostasis, and cell growth. [ABSTRACT FROM AUTHOR]

Fasting can be divided into intermittent short-term fasting (ISTF; frequent fasts lasting <24 h) and long-term fasting (LTF; fasts lasting >24 h and up to several weeks). ISTF extends lifespan and ameliorates a wide range of diseases in animal models. ISTF and ketogenic diets (KDs) reduce oxidative stress by direct actions on the mitochondria and by inducing the expression of antioxidant enzymes. The ketone body β-hydroxybutyrate suppresses inflammation by inhibiting the nucleotide oligomerization domain-like receptor family, pyrin domain-containing 3 (NLRP3) inflammasome. Human clinical trials demonstrated disease-modifying effects of ISTF in obesity, diabetes, and chronic inflammatory disorders. Frequent switching between ketogenic and nonketogenic states may optimize health and resilience. Intermittent short-term fasting (ISTF) and ketogenic diets (KDs) exert overlapping but not identical effects on cell metabolism, function, and resilience. Whereas health benefits of KD are largely mediated by the ketone bodies (KBs), ISTF engages additional adaptive physiological responses. KDs act mainly through inhibition of histone deacetylases (HDACs), reduction of oxidative stress, improvement of mitochondria efficiency, and control of inflammation. Mechanisms of action of ISTF include stimulation of autophagy, increased insulin and leptin sensitivity, activation of AMP-activated protein kinase (AMPK), inhibition of the mechanistic target of rapamycin (mTOR) pathway, bolstering mitochondrial resilience, and suppression of oxidative stress and inflammation. Frequent switching between ketogenic and nonketogenic states may optimize health by increasing stress resistance, while also enhancing cell plasticity and functionality. Intermittent short-term fasting (ISTF) and ketogenic diets (KDs) exert overlapping but not identical effects on cell metabolism, function, and resilience. Whereas health benefits of KD are largely mediated by the ketone bodies (KBs), ISTF engages additional adaptive physiological responses. KDs act mainly through inhibition of histone deacetylases, reduction of oxidative stress, improvement of mitochondria efficiency, and control of inflammation. Mechanisms of action of ISTF include stimulation of autophagy, increased insulin and leptin sensitivity, activation of AMP-activated protein kinase (AMPK), inhibition of the mechanistic target of rapamycin (mTOR) pathway, bolstering mitochondrial resilience, and suppression of oxidative stress and inflammation. Frequent switching between ketogenic and nonketogenic states may optimize health by increasing stress resistance, while also enhancing cell plasticity and functionality. [ABSTRACT FROM AUTHOR]

Alzheimer's disease (AD) is a leading cause of dementia in the elderly. Mitogen-activated protein kinase phosphatase 1 (MKP-1) plays a neuroprotective role in AD. However, the molecular mechanisms underlying the effects of MKP-1 on AD have not been extensively studied. MicroRNAs (miRNAs) regulate gene expression at the post-transcriptional level, thereby repressing mRNA translation. Here, we reported that the microRNA-429-3p (miR-429-3p) was significantly increased in the brain of APP23/PS45 AD model mice and N2AAPP AD model cells. We further found that miR-429-3p could downregulate MKP-1 expression by directly binding to its 3′-untranslated region (3′ UTR). Inhibition of miR-429-3p by its antagomir (A-miR-429) restored the expression of MKP-1 to a control level and consequently reduced the amyloidogenic processing of APP and A β accumulation. More importantly, intranasal administration of A-miR-429 successfully ameliorated the deficits of hippocampal CA1 long-term potentiation and spatial learning and memory in AD model mice by suppressing extracellular signal-regulated kinase (ERK1/2)-mediated GluA1 hyperphosphorylation at Ser831 site, thereby increasing the surface expression of GluA1-containing α -amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid receptors (AMPARs). Together, these results demonstrate that inhibiting miR-429-3p to upregulate MKP-1 effectively improves cognitive and synaptic functions in AD model mice, suggesting that miR-429 /MKP-1 pathway may be a novel therapeutic target for AD treatment. miR-429-3p inhibits MKP-1 expression by directly binding to its 3′UTR, consequently initiating ERK1/2-mediated hyperphosphorylation of GluA1 at S831, which ultimately leads to synaptic and cognitive dysfunction in AD. [Display omitted] [ABSTRACT FROM AUTHOR]

Heart failure (HF) is a highly morbid syndrome that seriously affects the physical and mental health of patients and generates an enormous socio-economic burden. In addition to cardiac myocyte oxidative stress and apoptosis, which are considered mechanisms for the development of HF, alterations in cardiac energy metabolism and pathological autophagy also contribute to cardiac abnormalities and ultimately HF. Silent information regulator 1 (Sirt1) and adenosine monophosphate-activated protein kinase (AMPK) are nicotinamide adenine dinucleotide (NAD+)-dependent deacetylases and phosphorylated kinases, respectively. They play similar roles in regulating some pathological processes of the heart through regulating targets such as peroxisome proliferator-activated receptor γ coactivator 1α (PGC-1α), protein 38 mitogen-activated protein kinase (p38 MAPK), peroxisome proliferator-activated receptors (PPARs), and mammalian target of rapamycin (mTOR). We summarized the synergistic effects of Sirt1 and AMPK in the heart, and listed the traditional Chinese medicine (TCM) that exhibit cardioprotective properties by modulating the Sirt1/AMPK pathway, to provide a basis for the development of Sirt1/AMPK activators or inhibitors for the treatment of HF and other cardiovascular diseases (CVDs). [Display omitted] • Sirt1 and AMPK are essential targets for regulating HF. • Sirt1 and AMPK influence energy metabolism disorders, oxidative stress, autophagy disorders and excessive apoptosis in HF. • Many traditional Chinese medicines have been demonstrated to exert cardioprotective effect by regulating Sirt1/AMPK pathway. [ABSTRACT FROM AUTHOR]

Tumor necrosis factor α (TNFα), an inflammatory cytokine, induces lipolysis and increases circulating concentrations of free fatty acids. In addition, TNFα is the first adipokine produced by adipose tissue in obesity, contributing to obesity-associated metabolic disease. Given that benzyl isothiocyanate (BITC) is a well-known anti-inflammatory agent, we hypothesized that BITC can ameliorate TNFα-induced lipolysis and investigated the working mechanisms involved. We first challenged 3T3-L1 adipocytes with TNFα to induce lipolysis, which was confirmed by increased glycerol release, decreased protein expression of peroxisome proliferator-activated receptor γ (PPARγ) and perilipin 1 (PLIN1), and increased phosphorylation of ERK, protein kinase A (PKA), and hormone-sensitive lipase (HSL). However, inhibition of ERK or PKA significantly attenuated the lipolytic activity of TNFα. Meanwhile, pretreatment with BITC significantly ameliorated the lipolytic activity of TNFα; the TNFα-induced phosphorylation of ERK, PKA, and HSL; the TNFα-induced ubiquitination of PPARγ; the TNFα-induced decrease in PPARγ nuclear protein binding to PPAR response element; and the TNFα-induced decrease in PLIN1 protein expression. Our results indicate that BITC ameliorates TNFα-induced lipolysis by inhibiting the ERK/PKA/HSL signaling pathway, preventing PPARγ proteasomal degradation, and maintaining PLIN1 protein expression. BITC inhibits TNFα-induced ubiquitination of PPARγ and subsequent proteasomal degradation and PPARγ is a critical transcription factor for PLIN1, which acts as a barrier to the breakdown of storage lipids. In addition, TNFα activates the ERK/PKA/HSL signaling pathway participating in lipolysis, which is suppressed by BITC. Abbreviations: BITC, benzyl isothiocyanate; DG, diglyceride; FFA, free fatty acid; HSL, hormone-sensitive lipase; MG, monoglyceride; PKA, protein kinase A; PLIN1, perilipin 1; PPARγ, peroxisome proliferator-activated receptor γ; TG, triglyceride; TNFα, tumor necrosis factor α. [Display omitted] [ABSTRACT FROM AUTHOR]

The PAH1-encoded phosphatidate (PA) phosphatase is a major source of diacylglycerol for the production of the storage lipid triacylglycerol and a key regulator for the de novo phospholipid synthesis in Saccharomyces cerevisiae. The catalytic function of Pah1 depends on its membrane localization which is mediated through its phosphorylation by multiple protein kinases and dephosphorylation by the Nem1-Spo7 protein phosphatase complex. The full-length Pah1 is composed of a catalytic core (N-LIP and HAD-like domains, amphipathic helix, and the WRDPLVDID domain) and non-catalytic regulatory sequences (intrinsically disordered regions, RP domain, and acidic tail) for phosphorylation and interaction with Nem1-Spo7. How the catalytic core regulates Pah1 localization and cellular function is not clear. In this work, we analyzed a variant of Pah1 (i.e., Pah1-CC (catalytic core)) that is composed only of the catalytic core. Pah1-CC expressed on a low-copy plasmid complemented the pah1Δ mutant phenotypes (e.g., nuclear/ER membrane expansion, reduced levels of triacylglycerol, and lipid droplet formation) without requiring Nem1-Spo7. The cellular function of Pah1-CC was supported by its PA phosphatase activity mostly associated with the membrane fraction. Although functional, Pah1-CC was distinct from Pah1 in the protein and enzymological properties, which include overexpression toxicity, association with heat shock proteins, and significant reduction of the Vmax value. These findings on the Pah1 catalytic core enhance the understanding of its structural requirements for membrane localization and activity control. [ABSTRACT FROM AUTHOR]

Epimedin B (EB) is one of the main flavonoid ingredients present in Epimedium brevicornum Maxim., a traditional herb widely used in China. Our previous study showed that EB was a stronger inducer of melanogenesis and an activator of tyrosinase (TYR). However, the role of EB in melanogenesis and the mechanism underlying the regulation remain unclear. Herein, as an extension to our previous investigation, we provide comprehensive evidence of EB-induced pigmentation in vivo and in vitro and elucidate the melanogenesis mechanism by assessing its effects on the TYR family of proteins (TYRs) in terms of expression, activity, and stability. The results showed that EB increased TYRs expression through microphthalmia-associated transcription factor-mediated p-Akt (referred to as protein kinase B (PKB))/glycogen synthase kinase 3β (GSK3β)/β-catenin, p-p70 S6 kinase cascades, and protein 38 (p38)/mitogen-activated protein (MAP) kinase (MAPK) and extracellular regulated protein kinases (ERK)/MAPK pathways, after which EB increased the number of melanosomes and promoted their maturation for melanogenesis in melanoma cells and human primary melanocytes/skin tissues. Furthermore, EB exerted repigmentation by stimulating TYR activity in hydroquinone- and N -phenylthiourea-induced TYR inhibitive models, including melanoma cells, zebrafish, and mice. Finally, EB ameliorated monobenzone-induced depigmentation in vitro and in vivo through the enhancement of TYRs stability by inhibiting TYR misfolding, TYR-related protein 1 formation, and retention in the endoplasmic reticulum and then by downregulating the ubiquitination and proteolysis processes. These data conclude that EB can target TYRs and alter their expression, activity, and stability, thus stimulating their pigmentation function, which might provide a novel rational strategy for hypopigmentation treatment in the pharmaceutical and cosmetic industries. [Display omitted] • It is first time to investigate and determine the pharmacological activity of Epimedin B in pigment synthesis. • Multiple depigmented models including in vivo and in vitro are utilized to evaluate effect of Epimedin B in pigment synthesis. • Epimedin B can target TYRs in terms of three aspects (expression, activity, and stability) to exert pigmentation function. [ABSTRACT FROM AUTHOR]

The occurrence of benign prostate hyperplasia (BPH) was related to disrupted sex steroid hormones, and metformin (Met) had a clinical response to sex steroid hormone-related gynaecological disease. However, whether Met exerts an antiproliferative effect on BPH via sex steroid hormones remains unclear. Here, our clinical study showed that along with prostatic epithelial cell (PEC) proliferation, sex steroid hormones were dysregulated in the serum and prostate of BPH patients. As the major contributor to dysregulated sex steroid hormones, elevated dihydrotestosterone (DHT) had a significant positive relationship with the clinical characteristics of BPH patients. Activation of adenosine 5'-monophosphate (AMP)-activated protein kinase (AMPK) by Met restored dysregulated sex steroid hormone homeostasis and exerted antiproliferative effects against DHT-induced proliferation by inhibiting the formation of androgen receptor (AR)-mediated Yes-associated protein (YAP1)-TEA domain transcription factor (TEAD4) heterodimers. Met's anti-proliferative effects were blocked by AMPK inhibitor or YAP1 overexpression in DHT-cultured BPH-1 cells. Our findings indicated that Met would be a promising clinical therapeutic approach for BPH by inhibiting dysregulated steroid hormone-induced PEC proliferation. Target and molecular mechanism of Met improving sex steroid hormones-induced BPH. Met regained dysregulated sex steroid hormones homeostasis, and inhibited PECs proliferation induced by dysregulated sex steroid hormones in BPH, and the anti-proliferation effects of Met was resulted from the regulation effect of AR-mediated YAP1-TEAD4 complex. [Display omitted] • Sex steroid hormone compositions were dysregulated in the serum and prostate of BPH patients. • Metformin inhibited PEC proliferation of BPH rats and DHT-cultured BPH-1 cells. • Metformin suppressed DHT/AR-induced the formation of YAP1-TEAD4 complex. • YAP1-TEAD4 complex was involved in the anti-proliferative effect of metformin. [ABSTRACT FROM AUTHOR]