Protein Methylation Research Articles

M6a Methylation: A New Avenue for Control of the Development of Type 2 Diabetes Mellitus and its Complications.

The rapidly emerging prevalence of type 2 diabetes mellitus (T2DM) and its associated complications have formed an increasingly serious threat to human life and health. Therefore, there is an urgent requirement to investigate the pathogenesis of T2DM and its complications, which will be conducive to discovering effective drugs for prevention and treatment. N6-methyladenosine (m6A) methylation is the most abundant and prevalent epigenetic modification of mRNA in mammals. m6A methylation is a dynamically reversible epigenetic transcriptome modification process that is jointly regulated by methyltransferases, demethylases and methylated reading proteins, which control the fate of target mRNAs through influencing splicing, translation and decay. Recent studies have revealed that m6A methylation plays an important role in β cellular function, insulin sensitivity and glycolipid metabolism. In this review, we summarized the current roles of m6A methylation in T2DM and T2DM-related complications such as diabetes nephropathy (DN), diabetes cardiovascular disease (DCD) and diabetes retinopathy (DR). Additionally, we sought the potential mechanism of m6A in T2DM and related complications, which may provide a rationale and strategy for potential therapeutic targeting of T2DM and its complications.

Insights into E-Cadherin Impairment in CDH1-Unaltered Invasive Lobular Carcinoma: A Comprehensive Bioinformatic Study.

Invasive lobular carcinoma exhibits unique morphological features frequently associated with alterations in CDH1. Although some studies have identified abnormalities in adhesion factors other than E-cadherin, the molecular mechanisms underlying E-cadherin abnormalities in CDH1-unaltered invasive lobular carcinoma remain poorly understood. In this study, we investigated the molecular underpinnings of E-cadherin dysregulation in invasive lobular carcinoma in the absence of CDH1 gene alterations, using comprehensive bioinformatic analyses. We conducted a comparative study of CDH1-mutated and non-mutated invasive lobular carcinoma and evaluated the differences in mRNA levels, reverse-phase protein array, methylation, and miRNAs. We observed that invasive lobular carcinoma cases without CDH1 alterations exhibited a significantly higher incidence of the Claudin-low subtype (p < 0.01). The results of the reverse-phase protein array indicate no significant difference in E-cadherin expression between CDH1-mutated and non-mutated cases. Therefore, abnormalities in E-cadherin production also exist in CDH1 non-mutated invasive lobular carcinoma. Considering that there are no differences in mRNA levels and methylation status, post-translational modifications are the most plausible explanation for the same. Hence, future studies should focus on elucidating the mechanism underlying E-cadherin inactivation via post-translational modifications in CDH1 non-mutated invasive lobular carcinoma.

Poly-GR Impairs PRMT1-Mediated Arginine Methylation of Disease-Linked RNA-Binding Proteins by Acting as a Substrate Sink.

Dipeptide repeat proteins (DPRs) are aberrant protein species found in C9orf72-linked amyotrophic lateral sclerosis (ALS) and frontotemporal dementia (FTD), two neurodegenerative diseases characterized by the cytoplasmic mislocalization and aggregation of RNA-binding proteins (RBPs). In particular, arginine (R)-rich DPRs (poly-GR and poly-PR) have been suggested to promiscuously interact with multiple cellular proteins and thereby exert high cytotoxicity. Components of the protein arginine methylation machinery have been identified as modulators of DPR toxicity and/or potential cellular interactors of R-rich DPRs; however, the molecular details and consequences of such an interaction are currently not well understood. Here, we demonstrate that several members of the family of protein arginine methyltransferases (PRMTs) can directly interact with R-rich DPRs in vitro and in the cytosol. In vitro, R-rich DPRs reduce solubility and promote phase separation of PRMT1, the main enzyme responsible for asymmetric arginine-dimethylation (ADMA) in mammalian cells, in a concentration- and length-dependent manner. Moreover, we demonstrate that poly-GR interferes more efficiently than poly-PR with PRMT1-mediated arginine methylation of RBPs such as hnRNPA3. We additionally show by two alternative approaches that poly-GR itself is a substrate for PRMT1-mediated arginine dimethylation. We propose that poly-GR may act as a direct competitor for arginine methylation of cellular PRMT1 targets, such as disease-linked RBPs.

Supplementation of l-arginine in pregnant gilts affects the protein abundance of DNMT1 in 35-day fetuses

Maternal nutrition is one of the main environmental factors regulating gene expression during fetal development through epigenetic modifications. Some nutrients, such as the amino acid l-arginine, are added to maternal diets to modulate gene expression, improve the reproductive performance of females, and enhance conceptus development. This study investigated the hypothesis that supplementation of pregnant gilts with l-arginine regulates gene expression in conceptuses through epigenetic mechanisms. For this, fetal programming phenotypic markers, the expression of key epigenetic genes, and the abundance of DNA methylation proteins (DNMT3A and DNMT1) were evaluated in 25- and 35-day conceptuses from gilts supplemented (ARG) or not (CON) with 1.0 % l-arginine during early gestation. At 25 days, there were no significant differences in phenotypic markers between CON and ARG embryos (P > 0.05). Similarly, no differences were found between CON and ARG fetuses at 35 days (P > 0.05). Maternal supplementation with l-arginine did not influence the expression of the evaluated key epigenetic genes in pig embryos or fetuses, nor DNMT3A protein abundance (P > 0.05); on the other hand, DNMT1 protein abundance was lower in ARG fetuses (P = 0.002). It is concluded that supplementation of l-arginine in pregnant gilts affects epigenetic mechanisms, such as DNA methylation, in 35-day fetuses through regulation of DNMT1 levels. Further studies using transcriptomic and proteomic analysis could reveal additional epigenetic modifications in embryos and fetuses following maternal supplementation with l-arginine.

The modification role and tumor association with a methyltransferase: KMT2C.

Histone methylation can affect chromosome structure and binding to other proteins, depending on the type of amino acid being modified and the number of methyl groups added, this modification may promote transcription of genes (H3K4me2, H3K4me3, and H3K79me3) or reduce transcription of genes (H3K9me2, H3K9me3, H3K27me2, H3K27me3, and H4K20me3). In addition, advances in tumor immunotherapy have shown that histone methylation as a type of protein post-translational modification is also involved in the proliferation, activation and metabolic reprogramming of immune cells in the tumor microenvironment. These post-translational modifications of proteins play a crucial role in regulating immune escape from tumors and immunotherapy. Lysine methyltransferases are important components of the post-translational histone methylation modification pathway. Lysine methyltransferase 2C (KMT2C), also known as MLL3, is a member of the lysine methyltransferase family, which mediates the methylation modification of histone 3 lysine 4 (H3K4), participates in the methylation of many histone proteins, and regulates a number of signaling pathways such as EMT, p53, Myc, DNA damage repair and other pathways. Studies of KMT2C have found that it is aberrantly expressed in many diseases, mainly tumors and hematological disorders. It can also inhibit the onset and progression of these diseases. Therefore, KMT2C may serve as a promising target for tumor immunotherapy for certain diseases. Here, we provide an overview of the structure of KMT2C, disease mechanisms, and diseases associated with KMT2C, and discuss related challenges.

Abstract Mo122: Investigating the Role of the Lysine Methyltransferase SMYD1 in Striated Muscle Motor Domain Folding

Properly coordinated folding and assembly of myosin into the sarcomere thick filament is critical for cardiac and skeletal muscle development, function, growth, and maintenance. The motor domain of myosin heavy chain (MHC) requires muscle-specific chaperones for proper folding, making it currently impossible to fold in non-muscle cells. Previous studies found that HSP90 and UNC45B are necessary, but not sufficient, for folding the motor domain, indicating other factors are involved. Defining the minimal set of factors for motor domain folding would allow for development of an improved expression system and accelerate the study of myosin mutations and myosin-targeted drug development. We hypothesize that the muscle-specific lysine methyltransferase SMYD1 contributes to MHC folding and stability. We found SMYD1 associates with UNC45B, HSP90, and MHC in the adult human heart. To study motor domain folding we generated a human MYH7 motor domain fused to GFP (MD::GFP). Transfection into COS-7 cells resulted in unfolded aggregates while adenoviral infection of C 2 C 12 myotubes leads to folded and functional MD::GFP. Interestingly, the folded MD::GFP contains lysine trimethylation, which may arise from the methyltransferase activity of SMYD1. When COS-7 cells were co-transfected with UNC45B and SMYD1, we observed increased total protein levels of non-functional MD::GFP without affecting RNA levels as assessed by RT-qPCR. This effect was not observed when MD::GFP was co-transfected with UNC45B or SMYD1 alone and is dependent on HSP90. We found that UNC45B and SMYD1 stabilized MD::GFP as determined by cycloheximide chase assays. We also found SMYD1 associates with MD::GFP, suggesting this stabilizing effect is due to a direct interaction. Interestingly, a catalytically inactive SMYD1 also increased MD::GFP protein levels in COS-7 cells, consistent with our observation that no significant changes in global protein methylation were observed in the presence of WT SMYD1. Our data suggests that SMYD1 stabilizes the motor domain of striated muscle myosin in conjunction with UNC45B and HSP90, in a manner that is independent of its methyltransferase activity. However, UNC45B and SMYD1 are insufficient for motor domain folding in a non-muscle cell line, suggesting that additional folding factors or proper stoichiometric ratios of chaperones/co-chaperones may be required.

Protein Arginine Methyltransferases: Emerging Targets in Cardiovascular and Metabolic Disease.

Cardiovascular diseases (CVDs) and metabolic disorders stand as formidable challenges that significantly impact the clinical outcomes and living quality for afflicted individuals. An intricate comprehension of the underlying mechanisms is paramount for the development of efficacious therapeutic strategies. Protein arginine methyltransferases (PRMTs), a class of enzymes responsible for the precise regulation of protein methylation, have ascended to pivotal roles and emerged as crucial regulators within the intrinsic pathophysiology of these diseases. Herein, we review recent advancements in research elucidating on the multifaceted involvements of PRMTs in cardiovascular system and metabolic diseases, contributing significantly to deepen our understanding of the pathogenesis and progression of these maladies. In addition, this review provides a comprehensive analysis to unveil the distinctive roles of PRMTs across diverse cell types implicated in cardiovascular and metabolic disorders, which holds great potential to reveal novel therapeutic interventions targeting PRMTs, thus presenting promising perspectives to effectively address the substantial global burden imposed by CVDs and metabolic disorders.

N6-methyladenosine-mediated LINC01087 promotes lung adenocarcinoma progression by regulating miR-514a-3p to upregulatecentrosome protein 55.

Long noncoding RNAs are key players in the development of lung adenocarcinoma (LUAD). The present study elucidated the role of LINC01087 in LUAD development. Cell vitality and apoptosis were assessed by the CCK-8 assay and flow cytometry, respectively. The transwell assay was adopted to evaluate cell migration and invasion. Levels of m6A modification of LINC01087 were determined using the methylated RNA binding protein immunoprecipitation assay. The interactions among LINC01087, miR-514a-3p, and centrosome protein 55 (CEP55) were evaluated using dual-luciferase reporter, RNA immunoprecipitation, and RNA-RNA pull-down assays. LINC01087 was highly expressed in LUAD, and its downregulation restrained cancer cell proliferation, migration, invasion, and epithelial-mesenchymal transition in vitro as well as tumor growth in a xenograft tumor model. Overexpression of miR-514a-3p inhibited malignant phenotypes in LUAD cells by inactivating RhoA/ROCK1 signaling via the suppression of CEP55 expression. Mechanistically, RBM15 increased the expression and mRNA stability of LINC01087 by mediating its m6A modification and LINC01087 induced CEP55 expression by sponging miR-514a-3p. RBM15-induced LINC01087 upregulation accelerated LUAD progression by regulating the miR-514a-3p/CEP55/RhoA/ROCK1 axis, illustrating the potential of LINC01087 as a novel target for LUAD therapy.

DNA Methylation-derived biological age and long-term mortality risk in subjects with type 2 diabetes

BackgroundIndividuals with type 2 diabetes (T2D) face an increased mortality risk, not fully captured by canonical risk factors. Biological age estimation through DNA methylation (DNAm), i.e. the epigenetic clocks, is emerging as a possible tool to improve risk stratification for multiple outcomes. However, whether these tools predict mortality independently of canonical risk factors in subjects with T2D is unknown.MethodsAmong a cohort of 568 T2D patients followed for 16.8 years, we selected a subgroup of 50 subjects, 27 survived and 23 deceased at present, passing the quality check and balanced for all risk factors after propensity score matching. We analyzed DNAm from peripheral blood leukocytes using the Infinium Human MethylationEPIC BeadChip (Illumina) to evaluate biological aging through previously validated epigenetic clocks and assess the DNAm-estimated levels of selected inflammatory proteins and blood cell counts. We tested the associations of these estimates with mortality using two-stage residual-outcome regression analysis, creating a reference model on data from the group of survived patients. ResultsDeceased subjects had higher median epigenetic age expressed with DNAmPhenoAge algorithm (57.49 [54.72; 60.58] years. vs. 53.40 [49.73; 56.75] years; p = 0.012), and accelerated DunedinPoAm pace of aging (1.05 [1.02; 1.11] vs. 1.02 [0.98; 1.06]; p = 0.012). DNAm PhenoAge (HR 1.16, 95% CI 1.05–1.28; p = 0.004) and DunedinPoAm (HR 3.65, 95% CI 1.43–9.35; p = 0.007) showed an association with mortality independently of canonical risk factors. The epigenetic predictors of 3 chronic inflammation-related proteins, i.e. CXCL10, CXCL11 and enRAGE, C-reactive protein methylation risk score and DNAm-based estimates of exhausted CD8 + T cell counts were higher in deceased subjects when compared to survived.ConclusionsThese findings suggest that biological aging, as estimated through existing epigenetic tools, is associated with mortality risk in individuals with T2D, independently of common risk factors and that increased DNAm-surrogates of inflammatory protein levels characterize deceased T2D patients. Replication in larger cohorts is needed to assess the potential of this approach to refine mortality risk in T2D.

Silencing of FTO inhibits oxidative stress to relieve neuropathic pain by m6A modification of GPR177.

Neuropathic pain (NP) is a challenging health condition owing to its complex nature and associated multiple etiologies. The occurrence of NP involves the abnormal activity of neurons mediated by oxidative stress (OS). Previous research has demonstrated that m6A methylation plays a role in the regulatory pathway of NP. This study aimed to investigate the specific molecular pathways through which m6A methylation modifiers alleviate NP. For this purpose, an NO rat model was developed via spared nerve injury (SNI), followed by quantifying the animal's pain assessment via paw withdrawal threshold (PWT) and paw withdrawal latency (PWL). The OS in SNI rats was evaluated by measuring reactive oxygen species, superoxide dismutase, and catalase (CAT) in spinal cord tissues. Moreover, quantitative-real-time polymerase chain reaction and western blot analysis were employed for detecting fat mass and obesity-associated (FTO) and GPR177 levels, while m6A levels of GPR117 were analyzed via MeRIP. The results indicated an enhanced OS with highly expressed FTO in spinal cord tissue samples, where knocking down Fto effectively relieved NP and OS in SNI rats. Mechanistic investigations revealed that Fto-mediated reduction of Grp177 m6A modification was involved in the WNT5a/TRPV1 axis-mediated OS remission of NP. Moreover, in vitro experiment results indicated that YTHDF2 was an important m6A methylated reading protein for this process. Fto silencing leads to increased m6A methylation of Grp177 through a YTHDF2-dependent mechanism, resulting in decreased Grp177 stability and ultimately reducing NP in rats by OS suppression.

Divergent DNA methylation patterns and gene expression in MYC and CDKN2B in canine transmissible venereal tumors.

Canine transmissible venereal tumor (CTVT), a unique transmissible cancer in dogs, affects the external genitalia and potentially spreads to other parts of the body. While somatic mutations in oncogenic and tumor-suppressing genes are linked to CTVT development, the impact of DNA methylation, which affects gene expression, remains unclear. This study explored whether DNA methylation in the promoter regions of the MYC oncogene and CDKN2B tumor suppressor genes in CTVTs is associated with their expression, both at the gene and protein levels. To investigate promoter DNA methylation of MYC and CDKN2B in CTVTs, we analyzed frozen tissue samples from genital CTVT (GTVTs) and extragenital CTVT (ETVTs). Genomic DNA was extracted, bisulfite-treated, and analyzed using bisulfite polymerase chain reaction (PCR) and sequencing. The messenger RNA and protein of MYC and CDKN2B were also extracted and assessed by real-time PCR and Western blotting. Matching formalin-fixed, paraffin-embedded blocks were used for immunohistochemical staining to visualize protein distribution in GTVT and ETVT tissues. Although both GTVT and ETVT samples showed MYC promoter methylation, the extent of methylation differed significantly. GTVTs displayed a much higher degree of methylation, potentially explaining the more pronounced downregulation of MYC gene expression and reduction in c-MYC protein levels observed in GTVTs compared with ETVTs. Our data revealed a prevalent hypermethylation pattern in the CDKN2B promoter across both sample types. However, DNA methylation, which was expected to have a suppressive effect, did not correlate with gene/protein expression. GTVTs displayed high protein levels despite significantly reduced CDKN2B expression. Conversely, ETVTs maintained regular CDKN2B expression but exhibited reduced protein production, suggesting a complex interplay between methylation and expression in these tumors. MYC demonstrated a clear association between its promoter methylation status, gene expression, and protein levels; however, CDKN2B lacked this correlation, implying the involvement of methylation-independent regulatory mechanisms and highlighting the need for further investigation.

RMSxAI: arginine methylation sites prediction from protein sequences using machine learning algorithms and explainable artificial intelligence

Protein methylation is a vital regulator of many biological processes at the post-translational level, and accurate prediction of protein methylation sites is essential for research and drug discovery. In this paper, we present a new method, namely RMSxAI, to predict the arginine methylation sites from primary sequences using machine learning algorithms and describe the predictions using explainable artificial intelligence (XAI) techniques. Leveraging experimentally validated methylated and unmethylated protein sequences from diverse organisms, we deduced several sequence features, encompassing physicochemical properties, amino acid composition, and evolutionary insights. Our results show that the proposed RMSxAI can predict protein methylation sites with high accuracy, bringing the F1 score up to 0.88 and overall accuracy up to 88.4%. We use various XAI methods to explain the output results. These include key features, partial occupancy maps, and local variation models that provide insight into key features and interactions that lead to predictions. Overall, our approach is relevant to research and drug discovery, and our results demonstrate the potential of machine learning algorithms and XAI methods to provide accurate and meaningful prediction of arginine methylation sites.

Protein Arginine Methyltransferase CARM1 in Human Breast Cancer.

Coactivator-associated arginine methyltransferase 1 (CARM1) is a protein arginine methyltransferase that deposits asymmetrical dimethylation marks on both histone and nonhistone substrates. The regulatory role of CARM1 in transcription was first identified in estrogen receptor positive (ER+) breast cancer. Since then, the mechanism of CARM1 in activating ER-target genes has been further interrogated. CARM1 is expressed at the highest level in ER negative (ER-) breast cancer and higher expression correlates with poor prognosis, suggesting an oncogenic role of CARM1. Indeed, in ER- breast cancer, CARM1 can promote proliferation and metastasis at least partly through methylation of proteins and activation of oncogenes. In this review, we summarize the mechanisms of transcriptional activation by CARM1 in breast cancer. The methyltransferase activity of CARM1 is important for many of its functions; here, we also highlight the nonenzymatic roles of CARM1. We also cover the biological processes regulated by CARM1 that are often deregulated in cancer and the ways to harness CARM1 in cancer treatment.

Discovery of NSD2 non-histone substrates and design of a super-substrate

The human protein lysine methyltransferase NSD2 catalyzes dimethylation at H3K36. It has very important roles in development and disease but many mechanistic features and its full spectrum of substrate proteins are unclear. Using peptide SPOT array methylation assays, we investigate the substrate sequence specificity of NSD2 and discover strong readout of residues between G33 (-3) and P38 (+2) on H3K36. Unexpectedly, we observe that amino acid residues different from natural ones in H3K36 are preferred at some positions. Combining four preferred residues led to the development of a super-substrate which is methylated much faster by NSD2 at peptide and protein level. Molecular dynamics simulations demonstrate that this activity increase is caused by distinct hyperactive conformations of the enzyme-peptide complex. To investigate the substrate spectrum of NSD2, we conducted a proteome wide search for nuclear proteins matching the specificity profile and discovered 22 peptide substrates of NSD2. In protein methylation studies, we identify K1033 of ATRX and K819 of FANCM as NSD2 methylation sites and also demonstrate their methylation in human cells. Both these proteins have important roles in DNA repair strengthening the connection of NSD2 and H3K36 methylation to DNA repair.

Dysregulation of arginine methylation in tumorigenesis.

Protein methylation, similar to DNA methylation, primarily involves post-translational modification (PTM) targeting residues of nitrogen-containing side-chains and other residues. Protein arginine methylation, occurred on arginine residue, is mainly mediated by protein arginine methyltransferases (PRMTs), which are ubiquitously present in a multitude of organisms and are intricately involved in the regulation of numerous biological processes. Specifically, PRMTs are pivotal in the process of gene transcription regulation, and protein function modulation. Abnormal arginine methylation, particularly in histones, can induce dysregulation of gene expression, thereby leading to the development of cancer. The recent advancements in modification mediated by PRMTs and cancer research have had a profound impact on our understanding of the abnormal modification involved in carcinogenesis and progression. This review will provide a defined overview of these recent progression, with the aim of augmenting our knowledge on the role of PRMTs in progression and their potential application in cancer therapy.

FBXO7 ubiquitinates PRMT1 to suppress serine synthesis and tumor growth in hepatocellular carcinoma

Cancer cells are often addicted to serine synthesis to support growth. How serine synthesis is regulated in cancer is not well understood. We recently demonstrated protein arginine methyltransferase 1 (PRMT1) is upregulated in hepatocellular carcinoma (HCC) to methylate and activate phosphoglycerate dehydrogenase (PHGDH), thereby promoting serine synthesis. However, the mechanisms underlying PRMT1 upregulation and regulation of PRMT1-PHGDH axis remain unclear. Here, we show the E3 ubiquitin ligase F-box-only protein 7 (FBXO7) inhibits serine synthesis in HCC by binding PRMT1, inducing lysine 37 ubiquitination, and promoting proteosomal degradation of PRMT1. FBXO7-mediated PRMT1 downregulation cripples PHGDH arginine methylation and activation, resulting in impaired serine synthesis, accumulation of reactive oxygen species (ROS), and inhibition of HCC cell growth. Notably, FBXO7 is significantly downregulated in human HCC tissues, and inversely associated with PRMT1 protein and PHGDH methylation level. Overall, our study provides mechanistic insights into the regulation of cancer serine synthesis by FBXO7-PRMT1-PHGDH axis, and will facilitate the development of serine-targeting strategies for cancer therapy.

M2 macrophages regulate KDM6B/PFKFB2 metabolic reprogramming of cervical squamous cell carcinoma through CXCL1.

Macrophages in the tumor microenvironment can polarize into M1 or M2 forms, with M2 macrophages (M2φ) promoting tumor growth and metastasis in cervical squamous cell carcinoma (CESC). This study explored the effects of M2φ on CESC metabolic reprogramming both in vitro and in vivo. Results showed that M2φ secreted CXCL1, which significantly increased CESC migration and metabolic regulation. Further experiments revealed that CXCL1 upregulated KDM6B to enhance PFKFB2 transcriptional activity, thus regulating CESC glucose metabolism. Transcriptome sequencing screened 5 upregulated genes related to glycolysis, with PFKFB2 showing the most significant increase in cells treated with rCXCL1. Dual-luciferase reporter assay confirmed that rCXCL1 enhances PFKFB2 transcriptional activity. Bioinformatics analysis revealed a high correlation between expressions of KDM6B and PFKFB2 in CESC. Mechanistic experiments demonstrated that KDM6B inhibited H3K27me3 modification to activate PFKFB2 transcriptional expression. In conclusion, M2φ secreted CXCL1 to promote CESC cell migration and invasion, and CXCL1 activated KDM6B expression in CESC cells, inhibiting H3K27 protein methylation modification, and enhanced PFKFB2 transcriptional activity to regulate CESC glucose metabolism. These results provided new insights into the complex interplay between the immune system and cancer metabolism, which may have broader implications for understanding and treating other types of cancer.

Increased FGF2 expression promotes immune cell infiltration and correlates with an unfavorable prognosis in thyroid cancer

To delve into the expression and functions of FGF2 in patient with thyroid cancer (THCA), we conducted a systematic analysis of the association of FGF2 with immune cell infiltration, and prognosis in THCA patients. The transcription and protein levels, methylation, and biological properties of FGF2 were examined, along with its correlation with prognosis and immune cell infiltration in THCA patients using online databases UALCAN, Human Protein Atlas, Kaplan-Meier Plotter, DNMIVD, cBioPortal, GEPIA, Metascape, Linkedomics and TIMER. Clinical samples were collected for Western blot analyses. FGF2 was substantially overexpressed in the tumor group and shown correlations with age, tumor histology, nodal metastasis, and cancer stages. Moreover, higher expression of FGF2 (HR = 3.42, 95 % CI:1.57-7.44, p = 0.00099) was greatly correlated with poorer relapse-free survival in THCA patients, particularly in female patients. FGF2 methylation level was increased in the tumor group (p = 1.29E-6), and higher methylation levels of FGF2 were positively correlated with the poorer progression-free interval in THCA patients (p = 0.015). FGF2 mutations were markedly associated with shorter disease-free survival, with a mutation rate of 6 % among the total 498 THCA patients. FGF2 functions were potentially related to cell adhesion, cytokine-cytokine receptor interaction and angiogenesis. FGF2 expression showed positive correlations with the infiltration of B cells (Cor = 0.569, p = 1.04e-42), CD4+ T cells (Cor = 0.555, p = 9.43e-41), macrophages (Cor = 0.438, p = 2.94e-42), neutrophils (Cor = 0.578, p = 9.354e-45), and dendritic cells (Cor = 0.591, p = 5.00e-47). FGF2 is a potential prognostic marker in THCA patients, with its functions possibly related to cell adhesion, interaction of the cytokine-cytokine receptor, angiogenesis, and the promotion of multiple immune cell infiltration.

Dual inhibition of AKT and ERK1/2 pathways restores the expression of progesterone Receptor-B in endometriotic lesions through epigenetic mechanisms

Endometriosis is an estrogen-dependent and progesterone-resistant gynecological inflammatory disease of reproductive-age women. Progesterone resistance, loss of progesterone receptor -B (PR-B) in the stromal cells of the endometrium, is one of the hallmarks of endometriosis and a major contributing factor for infertility in endometriosis patients. Loss of PR-B in the stromal cells of the endometriotic lesions poses resistance to the success of progesterone-based therapy. The working hypothesis is that PR-B is hypermethylated and epigenetically silenced, and inhibition of AKT and ERK1/2 pathways will decrease the hypermethylation, reverse the epigenetic silencing, and restore the expression of PR-B via DNA methylation and histone modification mechanisms in the endometriotic lesions. The objectives are to (i) determine the effects of dual inhibition of AKT and ERK1/2 pathways on the expression of PR-B and DNA methylation and histone modification protein machinery in the endometriotic lesions and (ii) identify the underlying epigenetic mechanisms of PR-B restoration in the endometriotic lesions. The results indicate that dual inhibition of AKT and ERK1/2 pathways decreases the hypermethylation, reverses the epigenetic silencing, and restores the expression of PR-B via DNA methylation and H3K9 and H3K27 methylation mechanisms in the endometriotic lesions or endometriotic stromal cells of human origin. These results support the novel concept that restored expression of PR-B in the endometriotic lesions and endometrium may improve the clinical outcome of progesterone therapy in endometriosis patients.

Characterization of the skeletal muscle arginine methylome in health and disease reveals remodeling in amyotrophic lateral sclerosis.

Arginine methylation is a protein posttranslational modification important for the development of skeletal muscle mass and function. Despite this, our understanding of the regulation of arginine methylation under settings of health and disease remains largely undefined. Here, we investigated the regulation of arginine methylation in skeletal muscles in response to exercise and hypertrophic growth, and in diseases involving metabolic dysfunction and atrophy. We report a limited regulation of arginine methylation under physiological settings that promote muscle health, such as during growth and acute exercise, nor in disease models of insulin resistance. In contrast, we saw a significant remodeling of asymmetric dimethylation in models of atrophy characterized by the loss of innervation, including in muscle biopsies from patients with myotrophic lateral sclerosis (ALS). Mass spectrometry-based quantification of the proteome and asymmetric arginine dimethylome of skeletal muscle from individuals with ALS revealed the largest compendium of protein changes with the identification of 793 regulated proteins, and novel site-specific changes in asymmetric dimethyl arginine (aDMA) of key sarcomeric and cytoskeletal proteins. Finally, we show that invivo overexpression of PRMT1 and aDMA resulted in increased fatigue resistance and functional recovery in mice. Our study provides evidence for asymmetric dimethylation as a regulator of muscle pathophysiology and presents a valuable proteomics resource and rationale for numerous methylated and nonmethylated proteins, including PRMT1, to be pursued for therapeutic development in ALS.