Post-translational Modifications Research Articles

Strain-dependent glutathionylation of fibronectin fibers impacts mechano-chemical behavior and primes an integrin switch

The extracellular matrix (ECM) is a protein polymer network that physically supports cells within a tissue. It acts as an important physical and biochemical stimulus directing cell behaviors. For fibronectin (Fn), a predominant component of the ECM, these physical and biochemical activities are inextricably linked as physical forces trigger conformational changes that impact its biochemical activity. Here, we analyze whether oxidative post-translational modifications, specifically glutathionylation, alter Fn’s mechano-chemical characteristics through stretch-dependent protein modification. ECM post-translational modifications represent a potential for time- or stimulus-dependent changes in ECM structure-function relationships that could persist over time with potentially significant impacts on cell and tissue behaviors. In this study, we show evidence that glutathionylation of Fn ECM fibers is stretch-dependent and alters Fn fiber mechanical properties with implications on the selectivity of engaging integrin receptors. These data demonstrate the existence of multimodal post-translational modification mechanisms within the ECM with high relevance to the microenvironmental regulation of downstream cell behaviors.

Characterization of monoclonal antibody charge variants under near-native separation conditions using nanoflow sheath liquid capillary electrophoresis-mass spectrometry

BackgroundMonoclonal antibodies (mAbs) undergo multiple post-translational modifications (PTMs) during production and storage, resulting for instance in charge and oxidized variants. PTMs need to be assessed as critical quality attributes to assure protein quality and safety. Capillary zone electrophoresis (CZE) enables efficient charge-based separation. The CZE method developed by He et al. (2011) is currently applied routinely in the pharmaceutical industry for profiling charge heterogeneity of mAbs. However, as the method relies on a non-volatile background electrolyte (BGE), it cannot be directly hyphenated with mass spectrometry (MS), hampering the identification of separated charge variants. ResultsThis study presents a CZE-UV/MS method using a neutral static capillary coating of hydroxypropyl methylcellulose combined with a volatile BGE at pH 5.0 to allow for MS-compatible mAb charge variant separations. The effect of several parameters, including pH and concentration of the BGE, applied voltage, and injected mAb concentrations on separation performance was investigated using a panel of commercially available mAbs. The optimized method was evaluated with IgG1 and IgG4 mAbs of varying pI (7.4–9.2) and degrees of heterogeneity. Basic and acidic variants were separated from the parent mAb using a BGE of 50 mM acetic acid adjusted to pH 5.0 with ammonium hydroxide. The relative abundances of charge variants determined with the new method showed a good correlation with the corresponding relative levels obtained with the method of He et al. CZE-MS coupling was accomplished using the nanoCEasy, a low-flow sheath liquid interface, which enabled the identification and quantitation of basic, acidic, and incomplete pyroglutamate variants, and glycoforms of the tested mAbs. SignificanceThis manuscript describes a new CZE-MS method that permits heterogeneity assessment of mAbs under MS-compatible conditions, providing charge variant separation.

Chemokine Receptor N-Terminus Charge Dictates Reliance on Post-Translational Modifications for Effective Ligand Capture and Following Boosting by Defense Peptides.

The chemokine receptors CCR1 and CCR5 display overlapping expression patterns and ligand dependency. Here we find that ligand activation of CCR5, not CCR1, is dependent on N-terminal receptor O-glycosylation. Release from O-glycosylation dependency is obtained by increasing CCR5 N-terminus acidity to the level of CCR1. Ligand activation of CCR5, not CCR1, drastically improves in the absence of glycosaminoglycans (GAGs). Ligand activity at both CCR1 and CCR5 is boosted by positively charged/basic peptides shown to interact with acidic chemokine receptor N-termini. We propose that receptors with an inherent low N-terminus acidity rely on post-translational modifications (PTMs) to efficiently compete with acidic entities in the local environment for ligand capture. Although crucial for initial ligand binding, strong electrostatic interactions between the ligand and the receptor N-terminus may counteract following insertion of the ligand into the receptor binding pocket and activation, a process that seems to be aided in the presence of basic peptides. Basic peptides bind to the naked CCR1 N-terminus, not the CCR5 N-terminus, explaining the loss of boosting of ligand-induced signaling via CCR5 in cells incapable of glycosylation.

Seasonal influence on miRNA expression dynamics of extracellular vesicles in equine follicular fluid.

Ovarian follicular fluid (FF) is a dynamic environment that changes with the seasons, affecting follicle development, ovulation, and oocyte quality. Cells in the follicles release tiny particles called extracellular vesicles (EVs) containing vital regulatory molecules, such as microRNAs (miRNAs). These miRNAs are pivotal in facilitating communication within the follicles through diverse signaling and information transfer forms. EV-coupled miRNA signaling is implicated to be associated with ovarian function, follicle and oocyte growth and response to various environmental insults. Herein, we investigated how seasonal variations directly influence the ovulatory and anovulatory states of ovarian follicles and how are they associated with follicular fluid EV-coupled miRNA dynamics in horses. Ultrasonographic monitoring and follicular fluid aspiration of preovulatory follicles in horses during the anovulatory (spring: non-breeding) and ovulatory (spring, summer, and fall: breeding) seasons and subsequent EV isolation and miRNA profiling identified significant variation in EV-miRNA cargo content. We identified 97 miRNAs with differential expression among the groups and specific clusters of miRNAs involved in the spring transition (miR-149, -200b, -206, -221, -328, and -615) and peak breeding period (including miR-143, -192, -451, -302b, -100, and let-7c). Bioinformatic analyses showed enrichments in various biological functions, e.g., transcription factor activity, transcription and transcription regulation, nucleic acid binding, sequence-specific DNA binding, p53 signaling, and post-translational modifications. Cluster analyses revealed distinct sets of significantly up- and down-regulated miRNAs associated with spring anovulatory (Cluster 1) and summer ovulation-the peak breeding season (Clusters 4 and 6). The findings from the current study shed light on the dynamics of FF-EV-coupled miRNAs in relation to equine ovulatory and anovulatory seasons, and their roles in understanding the mechanisms involved in seasonal shifts and ovulation during the breeding season warrant further investigation.

Activating the NFE2L1-ubiquitin-proteasome system by DDI2 protects from ferroptosis.

Ferroptosis is an iron-dependent, non-apoptotic form of cell death initiated by oxidative stress and lipid peroxidation. Recent evidence has linked ferroptosis to the action of the transcription factor Nuclear factor erythroid-2 derived,-like-1 (NFE2L1). NFE2L1 regulates proteasome abundance in an adaptive fashion, maintaining protein quality control to secure cellular homeostasis, but the regulation of NFE2L1 during ferroptosis and the role of the ubiquitin-proteasome system (UPS) herein are still unclear. In the present study, using an unbiased proteomic approach charting the specific ubiquitylation sites, we show that induction of ferroptosis leads to recalibration of the UPS. RSL3-induced ferroptosis inhibits proteasome activity and leads to global hyperubiquitylation, which is linked to NFE2L1 activation. As NFE2L1 resides in the endoplasmic reticulum tethered to the membrane, it undergoes complex posttranslational modification steps to become active and induce the expression of proteasome subunit genes. We show that proteolytic cleavage of NFE2L1 by the aspartyl protease DNA-damage inducible 1 homolog 2 (DDI2) is a critical step for the ferroptosis-induced feed-back loop of proteasome function. Cells lacking DDI2 cannot activate NFE2L1 in response to RSL3 and show global hyperubiquitylation. Genetic or chemical induction of ferroptosis in cells with a disrupted DDI2-NFE2L1 pathway diminishes proteasomal activity and promotes cell death. Also, treating cells with the clinical drug nelfinavir, which inhibits DDI2, sensitized cells to ferroptosis. In conclusion, our results provide new insight into the importance of the UPS in ferroptosis and highlight the role of the DDI2-NFE2L1 as a potential therapeutic target. Manipulating DDI2-NFE2L1 activity through chemical inhibition might help sensitizing cells to ferroptosis, thus enhancing existing cancer therapies.

Application of a risk score model based on glycosylation-related genes in the prognosis and treatment of patients with low-grade glioma.

Low-grade gliomas (LGG) represent a heterogeneous and complex group of brain tumors. Despite significant progress in understanding and managing these tumors, there are still many challenges that need to be addressed. Glycosylation, a common post-translational modification of proteins, plays a significant role in tumor transformation. Numerous studies have demonstrated a close relationship between glycosylation modifications and tumor progression. However, the biological function of glycosylation-related genes in LGG remains largely unexplored. Their potential roles within the LGG microenvironment are also not well understood. We collected RNA-seq data and scRNA-seq data from patients with LGG from TCGA and GEO databases. The glycosylation pathway activity scores of each cluster and each patient were calculated by irGSEA and GSVA algorithms, and the differential genes between the high and low glycosylation pathway activity score groups were identified. Prognostic risk profiles of glycosylation-related genes were constructed using univariate Cox and LASSO regression analyses and validated in the CGGA database. An 8 genes risk score signature including ASPM, CHI3L1, LILRA4, MSN, OCIAD2, PTGER4, SERPING1 and TNFRSF12A was constructed based on the analysis of glycosylation-related genes. Patients with LGG were divided into high risk and low risk groups according to the median risk score. Significant differences in immunological characteristics, TIDE scores, drug sensitivity, and immunotherapy response were observed between these groups. Additionally, survival analysis of clinical medication information in the TCGA cohort indicated that high risk and low risk groups have different sensitivities to drug therapy. The risk score characteristics can thus guide clinical medication decisions for LGG patients. Our study established glycosylation-related gene risk score signatures, providing new perspectives and approaches for prognostic prediction and treatment of LGG.

Nitric Oxide Signaling and Sensing in Age-Related Diseases.

Nitric oxide (NO) is a key signaling molecule involved in numerous physiological and pathological processes within the human body. This review specifically examines the involvement of NO in age-related diseases, focusing on the cardiovascular, nervous, and immune systems. The discussion delves into the mechanisms of NO signaling in these diseases, emphasizing the post-translational modifications of involved proteins, such as S-nitrosation and nitration. The review also covers the dual nature of NO, highlighting both its protective and harmful effects, determined by concentration, location, and timing. Additionally, potential therapies that modulate NO signaling, including the use of NO donors and nitric oxide synthases (NOSs) inhibitors in the treatment of cardiovascular, neurodegenerative, and oncological diseases, are analyzed. Particular attention is paid to the methods for the determination of NO and its derivatives in the context of illness diagnosis and monitoring. The review underscores the complexity and dual role of NO in maintaining cellular balance and suggests areas for future research in developing new therapeutic strategies.

Post-Translational Modifications of RNA-Modifying Proteins in Cellular Dynamics and Disease Progression.

RNA-modifying proteins, classified as "writers," "erasers," and "readers," dynamically modulate RNA by adding, removing, or interpreting chemical groups, thereby influencing RNA stability, functionality, and interactions. To date, over 170 distinct RNA chemical modifications and more than 100 RNA-modifying enzymes have been identified, with ongoing research expanding these numbers. Although significant progress has been made in understanding RNA modification, the regulatory mechanisms that govern RNA-modifying proteins themselves remain insufficiently explored. Post-translational modifications (PTMs) such as phosphorylation, ubiquitination, and acetylation are crucial in modulating the function and behavior of these proteins. However, the full extent of PTM influence on RNA-modifying proteins and their role in disease development remains to be fully elucidated. This review addresses these gaps by offering a comprehensive analysis of the roles PTMs play in regulating RNA-modifying proteins. Mechanistic insights are provided into how these modifications alter biological processes, contribute to cellular function, and drive disease progression. In addition, the current research landscape is examined, highlighting the therapeutic potential of targeting PTMs on RNA-modifying proteins for precision medicine. By advancing understanding of these regulatory networks, this review seeks to facilitate the development of more effective therapeutic strategies and inspire future research in the critical area of PTMs in RNA-modifying proteins.

USP24 promotes autophagy-dependent ferroptosis in hepatocellular carcinoma by reducing the K48-linked ubiquitination of Beclin1.

Ubiquitination is a post-translational modification (PTM), which is critical to maintain cell homeostasis. Ubiquitin-specific protease 24 (USP24) plays roles in various diseases, the mechanisms by which USP24 regulates hepatocellular carcinoma (HCC) remain poorly understood. In this study, USP24 is found to be significantly downregulated in HCC. Knocking down USP24 promotes HCC proliferation and migration, whereas USP24 overexpression inhibits HCC in vitro and in vivo. The endogenous interaction between USP24 and Beclin1 is confirmed. Mechanically, USP24 delays Beclin1 degradation by reducing its K48-linked ubiquitination, the effects of overexpressing USP24 on HCC proliferation can be partially reversed by silencing Beclin1. We find that increased autophagy is accompanied by ferroptosis in USP24 overexpressed HCC cells and USP24 increases the susceptibility of HCC to sorafenib. Collectively, this study highlights the critical role of USP24 in regulating autophagy-dependent ferroptosis by decreasing Beclin1 ubiquitination, suggesting that targeting USP24 may be a strategy for treating HCC.

Generation of dengue 3 envelope domain III using tobacco mosaic virus-based vector system and its immunological response mouse model by generating anti-dengue virus antibodies.

Producing recombinant proteins in plants has become a valuable alternative to traditional microbial or mammalian systems due to its cost-effectiveness, scalability, and ability to perform post-translational modifications. This study investigates the use of the Tobacco Mosaic Virus (TMV)-based vector system for producing the Dengue virus serotype 3 (DENV-3) envelope domain III (EDIII) protein in plants.. A fragment of the gene that encodes domain III of the dengue 3 envelope protein (D3EIII, comprising 300-420 amino acids), was effectively expressed within Nicotiana tabacum plants utilizing a transient expression system based on tobacco mosaic virus (TMV). The N-terminal 5' UTR region upstream of D3EIII notably enhanced protein yield in infected tissues. The produced recombinant protein exhibited reactivity with both (anti) D3EIII polyclonal antibodies and antibodies of anti-His tag. Upon injection of EDIII in mice, it stimulated the generation of antibodies against the dengue-specific virus. The induced antibodies demonstrated neutralizing activity against dengue virus type 3. These findings indicate that the TMV expression system is effective for producing dengue virus antigens in plants, resulting in antigens with appropriate properties and strong immunogenic potential.

Histone Tail Cleavage as a Mechanism for Epigenetic Regulation.

Histones are essential for DNA packaging and undergo post-translational modifications that significantly influence gene regulation. Among these modifications, histone tail cleavage has recently garnered attention despite being less explored. Cleavage by various proteases impacts processes such as stem cell differentiation, aging, infection, and inflammation, though the mechanisms remain unclear. This review delves into recent insights on histone proteolytic cleavage and its epigenetic significance, highlighting how chromatin, which serves as a dynamic scaffold, responds to signals through histone modification, replacement, and ATP-dependent remodeling. Specifically, histone tail cleavage is linked to critical cellular processes such as granulocyte differentiation, viral infection, aging, yeast sporulation, and cancer development. Although the exact mechanisms connecting histone cleavage to gene expression are still emerging, it is clear that this process represents a novel epigenetic transcriptional mechanism intertwined with chromatin dynamics. This review explores known histone tail cleavage events, the proteolytic enzymes involved, their impact on gene expression, and future research directions in this evolving field.

Integrating N -glycan and CODEX imaging reveal cell-specific protein glycosylation in healthy human lung.

N -linked glycosylation, the major post-translational modification of cellular proteins, is important for proper lung functioning, serving to fold, traffic, and stabilize protein structures and to mediate various cell-cell recognition events. Identifying cell-specific N -glycan structures in human lungs is critical for understanding the chemistry and mechanisms that guide cell-cell and cell-matrix interactions and determining nuanced functions of specific N -glycosylation. Our study, which used matrix-assisted laser desorption/ionization (MALDI) mass spectrometry imaging (MSI) combined with co-detection by indexing (CODEX) to reveal the cellular origin of N -glycans, is a significant step in this direction. This innovative technological combination enabled us to detect and differentiate N -glycans located in the vicinity of cells surrounding airways and blood vessels, parenchyma, submucosal glands, cartilage, and smooth muscles. The potential impact of our findings on future research is immense. For instance, our algorithm for grouping N -glycans based on their functional chemical features, combined with identifying group niches, paves the way for targeted studies. We found that fucosylated N -glycans are dominant around immune cells, tetra antennary N -glycans in the cartilage, high-mannose N -glycans surrounding the bronchus originate from associated collagenous structures, complex fucosylated-tetra antennary-polylactosamine N -glycans are spread over smooth muscle structures and in epithelial cells surrounding arteries, and N -glycans with Hex:6 HexNAc:6 compositions, which, according to our algorithm, can be ascribed to either tetra antennary or bisecting N -glycan, are highly abundant in the parenchyma. The findings suggest cell or region-specific functions for these localized glycan structures.

The O-GlcNAc database: introducing new features and tools developed from community feedback.

O-GlcNAc is a reversible post-translational modification found on serine and threonine residues of nucleocytoplasmic proteins. Four years ago, we released the O-GlcNAc Database( oglcnac.mcw.edu ), a comprehensive catalog of O-GlcNAcylated proteins that has become one of the most cited resources in the field, with hundreds of unique users per month. We are now presenting an updated O-GlcNAc Database, which includes nearly 20,000 O-GlcNAcylated proteins and 48 species, marking substantial growth in data volume and scope. This paper presents the most noteworthy features implemented over the last year, often originating from feedback from the O-GlcNAc community. Among these features, we provide a brief overview of the database content, introduce our new protein viewer mode, and discuss the implementation of subcellular localization information and its applications in the O-GlcNAc score. We also provide an interface to use CytOVS, a tool designed to evaluate and sort O-GlcNAcome datasets derived from MS experiments. In conclusion, this new and improved O-GlcNAc Database represents a significant advancement in providing a comprehensive and expanded resource for researchers in the field of O-GlcNAc biology.

Emerging roles of cancer-associated histone mutations in genomic instabilities.

Epigenetic mechanisms often fuel the quick evolution of cancer cells from normal cells. Mutations or aberrant expressions in the enzymes of DNA methylation, histone post-translational modifications, and chromatin remodellers have been extensively investigated in cancer pathogenesis; however, cancer-associated histone mutants have gained momentum in recent decades. Next-generation sequencing of cancer cells has identified somatic recurrent mutations in all the histones (H3, H4, H2A, H2B, and H1) with different frequencies for various tumour types. Importantly, the well-characterised H3K27M, H3G34R/V, and H3K36M mutations are termed as oncohistone mutants because of their wide roles, from defects in cellular differentiation, transcriptional dysregulation, and perturbed epigenomic profiles to genomic instabilities. Mechanistically, these histone mutants impart their effects on histone modifications and/or on irregular distributions of chromatin complexes. Recent studies have identified the crucial roles of the H3K27M and H3G34R/V mutants in the DNA damage response pathway, but their impacts on chemotherapy and tumour progression remain elusive. In this review, we summarise the recent developments in their functions toward genomic instabilities and tumour progression. Finally, we discuss how such a mechanistic understanding can be harnessed toward the potential treatment of tumours harbouring the H3K27M, H3G34R/V, and H3K36M mutations.

Histone lactylation drives CD8+ T cell metabolism and function.

The activation and functional differentiation of CD8+ T cells are linked to metabolic pathways that result in the production of lactate. Lactylation is a lactate-derived histone post-translational modification; however, the relevance of histone lactylation in the context of CD8+ T cell activation and function is not known. Here, we show the enrichment of H3K18 lactylation (H3K18la) and H3K9 lactylation (H3K9la) in human and mouse CD8+ T cells, which act as transcription initiators of key genes regulating CD8+ T cell function. Further, we note distinct patterns of H3K18la and H3K9la in CD8+ T cell subsets linked to their specific metabolic profiles. Additionally, we find that modulation of H3K18la and H3K9la by targeting metabolic and epigenetic pathways influence CD8+ T cell effector function, including antitumor immunity, in preclinical models. Overall, our study uncovers the potential roles of H3K18la and H3K9la in CD8+ T cells.

Exploring the dynamics and interactions of the N-myc transactivation domain through solution nuclear magnetic resonance spectroscopy.

Myc proteins are transcription factors crucial for cell proliferation. They have a C-terminal domain that mediates Max and DNA binding, and an N-terminal disordered region culminating in the transactivation domain (TAD). The TAD participates in many protein-protein interactions, notably with kinases that promote stability (Aurora-A) or degradation (ERK1, GSK3) via the ubiquitin-proteasome system. We probed the structure, dynamics and interactions of N-myc TAD using nuclear magnetic resonance (NMR) spectroscopy following its complete backbone assignment. Chemical shift analysis revealed that N-myc has two regions with clear helical propensity: Trp77-Glu86 and Ala122-Glu132. These regions also have more restricted ps-ns motions than the rest of the TAD, and, along with the phosphodegron, have comparatively high transverse (R2) 15N relaxation rates, indicative of slower timescale dynamics and/or chemical exchange. Collectively these features suggest differential propensities for structure and interaction, either internal or with binding partners, across the TAD. Solution studies on the interaction between N-myc and Aurora-A revealed a previously uncharacterised binding site. The specificity and kinetics of sequential phosphorylation of N-myc by ERK1 and GSK3 were characterised using NMR and resulted in no significant structural changes outside the phosphodegron. When the phosphodegron was doubly phosphorylated, N-myc formed a robust interaction with the Fbxw7-Skp1 complex, but mapping the interaction by NMR suggests a more extensive interface. Our study provides foundational insights into N-myc TAD dynamics and a backbone assignment that will underpin future work on the structure, dynamics, interactions and regulatory post-translational modifications of this key oncoprotein.

Inhibitors of Intracellular RyR2 Calcium Release Channels as Therapeutic Agents in Arrhythmogenic Heart Diseases.

Ryanodine receptor type 2 (RyR2) is the principal intracellular calcium release channel in the cardiac sarcoplasmic reticulum (SR). Pathological RyR2 hyperactivity generates arrhythmia risk in genetic and structural heart diseases. RYR2 gain-of-function mutations cause catecholaminergic polymorphic ventricular tachycardia. In structural heart diseases (i.e., heart failure), posttranslation modifications render RyR2 channels leaky, resulting in pathologic calcium release during diastole, contributing to arrhythmogenesis and contractile dysfunction. Hence, RyR2 represents a therapeutic target in arrhythmogenic heart diseases. We provide an overview of the structure and function of RyR2, and then review US Food and Drug Administration-approved and investigational RyR2 inhibitors. A therapeutic classification of RyR2 inhibitors is proposed based on their mechanism of action. Class I RyR2 inhibitors (e.g., flecainide) do not change SR calcium content and are primarily antiarrhythmic. Class II RyR2 inhibitors (e.g., dantrolene) increase SR calcium content, making them less effective as antiarrhythmics but preferable in conditions with reduced SR calcium content such as heart failure.

GBMPhos: A Gating Mechanism and Bi-GRU-Based Method for Identifying Phosphorylation Sites of SARS-CoV-2 Infection.

Phosphorylation, a reversible and widespread post-translational modification of proteins, is essential for numerous cellular processes. However, due to technical limitations, large-scale detection of phosphorylation sites, especially those infected by SARS-CoV-2, remains a challenging task. To address this gap, we propose a method called GBMPhos, a novel method that combines convolutional neural networks (CNNs) for extracting local features, gating mechanisms to selectively focus on relevant information, and a bi-directional gated recurrent unit (Bi-GRU) to capture long-range dependencies within protein sequences. GBMPhos leverages a comprehensive set of features, including sequence encoding, physicochemical properties, and structural information, to provide an in-depth analysis of phosphorylation sites. We conducted an extensive comparison of GBMPhos with traditional machine learning algorithms and state-of-the-art methods. Experimental results demonstrate the superiority of GBMPhos over existing methods. The visualization analysis further highlights its effectiveness and efficiency. Additionally, we have established a free web server platform to help researchers explore phosphorylation in SARS-CoV-2 infections. The source code of GBMPhos is publicly available on GitHub.

Dehydration priming remodels protein abundance and phosphorylation level regulating tolerance to subsequent dehydration or salt stress in creeping bentgrass

Dehydration priming (DP) induces stress memory which plays a positive role in plant adaptability, but it is not well understood how DP differentially regulates subsequent dehydration (cis priming) or salt (trans priming) tolerance at the post-translational level. Purpose of this study was to identify proteins, phosphorylation levels and sites, and relevant metabolic pathways for DP-induced dehydration or salt tolerance in Agrostis stolonifera. DP-induced differentially regulated proteins (DRPs) were mostly located in the cytoplasm, chloroplast, and cell membrane, and differentially regulated phosphoproteins (DRPPs) were mostly nuclear proteins and cytoplasmic proteins. DP regulated common phosphorylation sites ([SP] and [RxxS]) under dehydration and salt conditions and also individually affected 8 or 11 phosphorylation sites under dehydration or salt stress. DP-regulated DRPPs were mainly rich in glycolysis and glutathione metabolism pathways, RNA splicing, and dynamin family proteins under dehydration stress, whereas DP-regulated salt tolerance was mainly related to chlorophyll metabolism, photosynthesis, MAPK signaling cascade, and ABC transporter I family at the phosphorylation level. In addition, the DP also significantly up-regulated phosphorylation of histones (ATXR3 and SETD1A) in response to subsequent dehydration and salt stress as well as abundances of antioxidant enzymes, dynamin family protein, and KCS6 under dehydration stress or abundances of PETE, HMGA, XTH, and ABCI6 under salt stress, respectively. Transcriptomics analysis further indicated that DP-regulated dehydration or salt tolerance was also related to transcriptional regulation in the early stage. Current results provided better understanding of the role of stress memory in plant adaptability to repeated or crossed stress via post-translational modifications (PTMs). SignificanceRecurrent moderate drought may buffer drought legacies in many plant species. When plants were exposed to repeated drought stress, their adaptability to subsequent stress could be enhanced, which is known as “stress memory”. Dehydration priming has been found to be an important approach to induce stress memory. Current results provided better understanding of the role of stress memory in plant adaptability to repeated or crossed stress via post-translational modifications.

Desmoplakin CSM models unravel mechanisms regulating the binding to intermediate filaments and putative therapeutics for cardiocutaneous diseases.

Arrhythmogenic cardiomyopathy (AC) is a common cause of sudden cardiac arrest and death in young adults. It can be induced by different types of mutations throughout the desmoplakin gene including the R2834H mutation in the extreme carboxyterminus tail of desmoplakin (DP CT) which remains structurally uncharacterized and poorly understood. Here, we have created 3D models of DP CT which show the structural effects of AC-inducing mutations as well as the implications of post-translational modifications (PTMs). Our results suggest that, in absence of PTMs, positively charged wildtype DP CT likely folds back onto negatively-charged plectin repeat 14 of nearby plakin repeat domain C (PRD C) contributing to the recruitment of intermediate filaments (IFs). When phosphorylated and methylated, negatively-charged wildtype DP CT would then fold back onto positively-charged plectin repeat 17 of PRD C, promoting the repulsion of intermediate filaments. However, by preventing PTMs, the R2834H mutation would lead to the formation of a cytoplasmic mutant desmoplakin with a constitutively positive DP CT tail that would be aberrantly recruited by cytoplasmic IFs instead of desmosomes, potentially weakening cell-cell contacts and promoting AC. Virtual screening of FDA-approved drug libraries identified several promising drug candidates for the treatment of cardiocutaneous diseases through drug repurposing.