MicroRNA-721 regulates gluconeogenesis via KDM2A-mediated epigenetic modulation in diet-induced insulin resistance in C57BL/6J mice.

BACKGROUND: Aberrant gluconeogenesis is considered among primary drivers of hyperglycemia under insulin resistant conditions, with multiple studies pointing towards epigenetic dysregulation. Here we examine the role of miR-721 and effect of epigenetic modulator laccaic acid on the regulation of gluconeogenesis under high fat diet induced insulin resistance. RESULTS: Reanalysis of miRNA profiling data of high-fat diet-induced insulin-resistant mice model, GEO dataset (GSE94799) revealed a significant upregulation of miR-721, which was further validated in invivo insulin resistance in mice and invitro insulin resistance in Hepa 1-6 cells. Interestingly, miR-721 mimic increased glucose production in Hepa 1-6 cells via activation of FOXO1 regulated gluconeogenic program. Concomitantly, inhibition of miR-721 reduced glucose production in palmitate induced insulin resistant Hepa 1-6 cells by blunting the FOXO1 induced gluconeogenesis. Intriguingly, at epigenetic level, enrichment of the transcriptional activation mark H3K36me2 got decreased around the FOXO1 promoter. Additionally, identifying targets of miR-721 using miRDB.org showed H3K36me2 demethylase KDM2A as a potential target. Notably, miR-721 inhibitor enhanced KDM2A expression which correlated with H3K36me2 enrichment around FOXO1 promoter and the downstream activation of the gluconeogenic pathway. Furthermore, inhibition of miR-721 in high-fat diet-induced insulin-resistant mice resulted in restoration of KDM2A levels, concomitantly reducing FOXO1, PCK1, and G6PC expression, attenuating gluconeogenesis, hyperglycemia, and improving glucose tolerance. Interestingly, the epigenetic modulator laccaic acid also reduced the hepatic miR-721 expression and improved KDM2A expression, supporting our earlier report that laccaic acid attenuates insulin resistance by reducing gluconeogenesis. CONCLUSION: Our study unveils the role of miR-721 in regulating gluconeogenesis through KDM2A and FOXO1 under insulin resistance, pointing towards significant clinical and therapeutic implications for metabolic disorders. Moreover, the promising impact of laccaic acid highlights its potential as a valuable intervention in managing insulin resistance-associated metabolic diseases.

SKP2-mediated ubiquitination and degradation of KLF11 promotes osteoarthritis via modulation of JMJD3/NOTCH1 pathway.

Osteoarthritis (OA) is the main cause of cartilage damage and disability. This study explored the biological function of S-phase kinase-associated protein 2 (SKP2) and Kruppel-like factor 11 (KLF11) in OA progression and its underlying mechanisms. C28/I2 chondrocytes were stimulated with IL-1ß to mimic OA in vitro. We found that SKP2, Jumonji domain-containing protein D3 (JMJD3), and Notch receptor 1 (NOTCH1) were upregulated, while KLF11 was downregulated in IL-1ß-stimulated chondrocytes. SKP2/JMJD3 silencing or KLF11 overexpression repressed apoptosis and extracellular matrix (ECM) degradation in chondrocytes. Mechanistically, SKP2 triggered the ubiquitination and degradation of KLF11 to transcriptionally activate JMJD3, which resulted in activation of NOTCH1 through inhibiting H3K27me3. What's more, the in vivo study found that KLF11 overexpression delayed OA development in rats via restraining apoptosis and maintaining the balance of ECM metabolism. Taken together, ubiquitination and degradation of KLF11 regulated by SKP2 contributed to OA progression by activation of JMJD3/NOTCH1 pathway. Our findings provide promising therapeutic targets for OA.

JMJD6 Autoantibodies as a Potential Biomarker for Inflammation-Related Diseases.

Inflammation is closely associated with cerebrovascular diseases, cardiovascular diseases, diabetes, and cancers, and it is accompanied by the development of autoantibodies in the early stage of inflammation-related diseases. Hence, it is meaningful to discover novel antibody biomarkers targeting inflammation-related diseases. In this study, Jumonji C-domain-containing 6 (JMJD6) was identified by the serological identification of antigens through recombinant cDNA expression cloning. In particular, JMJD6 is an antigen recognized in serum IgG from patients with unstable angina pectoris (a cardiovascular disease). Then, the serum antibody levels were examined using an amplified luminescent proximity homogeneous assay-linked immunosorbent assay and a purified recombinant JMJD6 protein as an antigen. We observed elevated levels of serum anti-JMJD6 antibodies (s-JMJD6-Abs) in patients with inflammation-related diseases such as ischemic stroke, acute myocardial infarction (AMI), diabetes mellitus (DM), and cancers (including esophageal cancer, EC; gastric cancer; lung cancer; and mammary cancer), compared with the levels in healthy donors. The s-JMJD6-Ab levels were closely associated with some inflammation indicators, such as C-reactive protein and intima-media thickness (an atherosclerosis index). A better postoperative survival status of patients with EC was observed in the JMJD6-Ab-positive group than in the negative group. An immunohistochemical analysis showed that JMJD6 was highly expressed in the inflamed mucosa of esophageal tissues, esophageal carcinoma tissues, and atherosclerotic plaques. Hence, JMJD6 autoantibodies may reflect inflammation, thereby serving as a potential biomarker for diagnosing specific inflammation-related diseases, including stroke, AMI, DM, and cancers, and for prediction of the prognosis in patients with EC.

LncRNA-PCat19 acts as a ceRNA of miR-378a-3p to facilitate microglia activation and accelerate chronic neuropathic pain in rats by promoting KDM3A-mediated BDNF demethylation.

The pathogenesis of neuropathic pain (NP) is complex, and there are various pathological processes. Previous studies have suggested that lncRNA PCAT19 is abnormally expressed in NP conduction and affects the occurrence and development of pain. The aim of this study is to analyze the role and mechanism of PCAT19 in NP induced by chronic compressive nerve injury (CCI) in mice. In this study, C57BL/6 mice were applied to establish the CCI model. sh-PCAT19 was intrathecally injected once a day for 5 consecutive days from the second day after surgery. We discovered that PCat19 level was gradually up-regulated with the passage of modeling time. Downregulation of Iba-1-positive expression, M1/M2 ratio of microglia, and pro-inflammatory factors in the spinal cords of CCI-mice after PCat19 knock-downed was observed. Mechanically, the expression of miR-378a-3p was negatively correlated with KDM3A and PCat19. Deletion of KDM3A prevented H3K9me2 demethylation of BDNF promoter and suppressed BDNF expression. Further, KDM3A promotes CCI-induced neuroinflammation and microglia activation by mediating Brain-derived neurotrophic factor (BDNF) demethylation. Together, the results suggest that PCat19 may be involved in the development of NP and that PCat19 shRNA injection can attenuate microglia-induced neuroinflammation by blocking KDM3A-mediated demethylation of BDNF and BDNF release.

Co-targeting SKP2 and KDM5B inhibits prostate cancer progression by abrogating AKT signaling with induction of senescence and apoptosis.

BACKGROUND: Prostate cancer (PCa) is the second-leading cause of cancer mortalities in the United States and is the most commonly diagnosed malignancy in men. While androgen deprivation therapy (ADT) is the first-line treatment option to initial responses, most PCa patients invariably develop castration-resistant PCa (CRPC). Therefore, novel and effective treatment strategies are needed. The goal of this study was to evaluate the anticancer effects of the combination of two small molecule inhibitors, SZL-P1-41 (SKP2 inhibitor) and PBIT (KDM5B inhibitor), on PCa suppression and to delineate the underlying molecular mechanisms. METHODS: Human CRPC cell lines, C4-2B and PC3 cells, were treated with small molecular inhibitors alone or in combination, to assess effects on cell proliferation, migration, senescence, and apoptosis. RESULTS: SKP2 and KDM5B showed an inverse regulation at the translational level in PCa cells. Cells deficient in SKP2 showed an increase in KDM5B protein level, compared to that in cells expressing SKP2. By contrast, cells deficient in KDM5B showed an increase in SKP2 protein level, compared to that in cells with KDM5B intact. The stability of SKP2 protein was prolonged in KDM5B depleted cells as measured by cycloheximide chase assay. Cells deficient in KDM5B were more vulnerable to SKP2 inhibition, showing a twofold greater reduction in proliferation compared to cells with KDM5B intact (p < 0.05). More importantly, combined inhibition of KDM5B and SKP2 significantly decreased proliferation and migration of PCa cells as compared to untreated controls (p < 0.005). Mechanistically, combined inhibition of KDM5B and SKP2 in PCa cells abrogated AKT activation, resulting in an induction of both cellular senescence and apoptosis, which was measured via Western blot analysis and senescence-associated ß-galactosidase (SA-ß-Gal) staining. CONCLUSIONS: Combined inhibition of KDM5B and SKP2 was more effective at inhibiting proliferation and migration of CRPC cells, and this regimen would be an ideal therapeutic approach of controlling CRPC malignancy.

circ_JMJD1C expedites breast cancer progression by regulating miR-182-5p/JMJD1C/SOX4 axis.

Circular RNAs (circRNAs) are engaged in various types of cancers. This study aimed to investigate the roles of circ_0006743 (circ_JMJD1C) in breast cancer. The downstream of circ_JMJD1C and their interaction network was determined by bioinformatic analyses. Gene expression were analyzed through western blot and qRT-PCR assays. Functional assays were conducted in vitro and in vivo to verify circ_JMJD1C role in BC. FISH and confocal analysis indicated the cellular distribution of circ_JMJD1C. Luciferase reporter, RNA immune-precipitation (RIP) assays, as well as Pearson's correlation analysis, were implemented to test the relation of miR-182-5p, JMJD1C and circ_JMJD1C. Circ_JMJD1C and JMJD1C expression were both elevated, and their expression was positively correlated in BC. Circ_ JMJD1C knockdown hindered BC cell proliferation, invasion, and migration, along with epithelial-mesenchymal transition (EMT) in vitro and in vivo. Circ_JMJD1C facilitated BC progression by the miR-182-5p-JMJD1C axis. Circ_JMJD1C epigenetically upregulated SOX4. Circ_JMJD1C promotes the aggressiveness of BC via regulating miR-182-5p/JMJD1C/SOX4 axis. This may provide a novel and promising therapy targeting BC.

[The Regulatory Effect of RNA m6 A Methylation Modification on KDM4B Gene Expression in t (8;21) AML Cells by MeRIP-qPCR].

OBJECTIVE: To confirm the direct regulatory effect of WTAP-mediated RNA m6A modification on the KDM4B gene in t (8;21) acute myeloid leukemia (AML) cells through MeRIP combined with reverse transcription real-time quantitative PCR (RT-qPCR) technology. METHODS: The lentivirus-mediated shRNA target WTAP or KDM4B gene was used to transfect the t (8;21) AML cell lines: Kasumi-1 and SKNO-1, and cells transfected with randomly shuffled shRNA as the control. Using the Ultrapure RNA Extraction Kit (DNase I) to extract RNA. The Magna MeRIPTM m6A Kit was used to enrich methylated modified fragments, and detect the m6A methylated RNA regions by RT-qPCR, and the protein and mRNA expression levels of WTAP and KDM4B in cells were detected by Western blot and reverse transcription real-time quantitative PCR (RT-qPCR). Colony formation assays were used to detect the colony ability of cells in vitro. RESULTS: Silencing the expression of WTAP in Kasumi-1 cells, the enrichment of m6A methylation modification was significantly decreased in the 3'UTR of KDM4B mRNA(P < 0.01), and the protein(P < 0.001) and mRNA (Kasumi-1:P < 0.001; SKNO-1: P < 0.01) expression levels of KDM4B were also significantly inhibited in Kasumi-1 and SKNO-1 cells upon WTAP knockdown (all P < 0.01), accompanied by a significant decrease in the colony-forming ability of both cell lines (both P < 0.01). CONCLUSION: In t(8;21) AML cell lines, WTAP could regulate the expression of KDM4B by regulating the m6A modification of the 3'UTR of KDM4B mRNA, and silencing the expression of KDM4B could inhibit the cellular proliferation in vitro.

Targeting histone demethylases JMJD3 and UTX: selenium as a potential therapeutic agent for cervical cancer.

BACKGROUND: The intriguing connection between selenium and cancer resembles a captivating puzzle that keeps researchers engaged and curious. While selenium has shown promise in reducing cancer risks through supplementation, its interaction with epigenetics in cervical cancer remains a fascinating yet largely unexplored realm. Unraveling the intricacies of selenium's role and its interaction with epigenetic factors could unlock valuable insights in the battle against this complex disease. RESULT: Selenium has shown remarkable inhibitory effects on cervical cancer cells in various ways. In in vitro studies, it effectively inhibits the proliferation, migration, and invasion of cervical cancer cells, while promoting apoptosis. Selenium also demonstrates significant inhibitory effects on human cervical cancer-derived organoids. Furthermore, in an in vivo study, the administration of selenium dioxide solution effectively suppresses the growth of cervical cancer tumors in mice. One of the mechanisms behind selenium's inhibitory effects is its ability to inhibit histone demethylases, specifically JMJD3 and UTX. This inhibition is observed both in vitro and in vivo. Notably, when JMJD3 and UTX are inhibited with GSK-J4, similar biological effects are observed in both in vitro and in vivo models, effectively inhibiting organoid models derived from cervical cancer patients. Inhibiting JMJD3 and UTX also induces G2/M phase arrest, promotes cellular apoptosis, and reverses epithelial-mesenchymal transition (EMT). ChIP-qPCR analysis confirms that JMJD3 and UTX inhibition increases the recruitment of a specific histone modification, H3K27me3, to the transcription start sites (TSS) of target genes in cervical cancer cells (HeLa and SiHa cells). Furthermore, the expressions of JMJD3 and UTX are found to be significantly higher in cervical cancer tissues compared to adjacent normal cervical tissues, suggesting their potential as therapeutic targets. CONCLUSIONS: Our study highlights the significant inhibitory effects of selenium on the growth, migration, and invasion of cervical cancer cells, promoting apoptosis and displaying promising potential as a therapeutic agent. We identified the histone demethylases JMJD3 and UTX as specific targets of selenium, and their inhibition replicates the observed effects on cancer cell behavior. These findings suggest that JMJD3 and UTX could be valuable targets for selenium-based treatments of cervical cancer.

The Intellectual Disability Risk Gene Kdm5b Regulates Long-Term Memory Consolidation in the Hippocampus.

The histone lysine demethylase KDM5B is implicated in recessive intellectual disability disorders, and heterozygous, protein-truncating variants in KDM5B are associated with reduced cognitive function in the population. The KDM5 family of lysine demethylases has developmental and homeostatic functions in the brain, some of which appear to be independent of lysine demethylase activity. To determine the functions of KDM5B in hippocampus-dependent learning and memory, we first studied male and female mice homozygous for a Kdm5b Δ ARID allele that lacks demethylase activity. Kdm5b Δ ARID/ Δ ARID mice exhibited hyperactivity and long-term memory deficits in hippocampus-dependent learning tasks. The expression of immediate early, activity-dependent genes was downregulated in these mice and hyperactivated upon a learning stimulus compared with wild-type (WT) mice. A number of other learning-associated genes were also significantly dysregulated in the Kdm5b Δ ARID/ Δ ARID hippocampus. Next, we knocked down Kdm5b specifically in the adult, WT mouse hippocampus with shRNA. Kdm5b knockdown resulted in spontaneous seizures, hyperactivity, and hippocampus-dependent long-term memory and long-term potentiation deficits. These findings identify KDM5B as a critical regulator of gene expression and synaptic plasticity in the adult hippocampus and suggest that at least some of the cognitive phenotypes associated with KDM5B gene variants are caused by direct effects on memory consolidation mechanisms.

Glutamine-mediated epigenetic regulation of cFLIP underlies resistance to TRAIL in pancreatic cancer.

Tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) is a promising anticancer agent because it kills cancer cells while sparing normal cells. However, many cancers, including pancreatic ductal adenocarcinoma (PDAC), exhibit intrinsic or acquired resistance to TRAIL, and the molecular mechanisms underlying TRAIL resistance in cancers, particularly in PDAC, remain unclear. In this study, we demonstrated that glutamine (Gln) endows PDAC cells with resistance to TRAIL through KDM4C-mediated epigenetic regulation of cFLIP. Inhibition of glutaminolysis significantly reduced the cFLIP level, leading to TRAIL-mediated formation of death-inducing signaling complexes. Overexpression of cFLIP dramatically rescued PDAC cells from TRAIL/Gln deprivation-induced apoptosis. Alpha-Ketoglutarate (aKG) supplementation significantly reversed the decrease in the cFLIP level induced by glutaminolysis inhibition and rescued PDAC cells from TRAIL/Gln deprivation-induced apoptosis. Knockdown of glutamic-oxaloacetic transaminase 2, which facilitates the conversion of oxaloacetate and glutamate into aspartate and aKG, decreased aKG production and the cFLIP level and activated TRAIL-induced apoptosis. AKG-mediated epigenetic regulation was necessary for maintaining a high level of cFLIP. Glutaminolysis inhibition increased the abundance of H3K9me3 in the cFLIP promoter, indicating that Gln-derived aKG production is important for Jumonji-domain histone demethylase (JHDM)-mediated cFLIP regulation. The JHDM KDM4C regulated cFLIP expression by binding to its promoter, and KDM4C knockdown sensitized PDAC cells to TRAIL-induced apoptosis. The present findings suggest that Gln-derived aKG production is required for KDM4C-mediated epigenetic regulation of cFLIP, which leads to resistance to TRAIL.

Histone lysine demethylase 4 family proteins maintain the transcriptional program and adrenergic cellular state of MYCN-amplified neuroblastoma.

Neuroblastoma with MYCN amplification (MNA) is a high-risk disease that has a poor survival rate. Neuroblastoma displays cellular heterogeneity, including more differentiated (adrenergic) and more primitive (mesenchymal) cellular states. Here, we demonstrate that MYCN oncoprotein promotes a cellular state switch in mesenchymal cells to an adrenergic state, accompanied by induction of histone lysine demethylase 4 family members (KDM4A-C) that act in concert to control the expression of MYCN and adrenergic core regulatory circulatory (CRC) transcription factors. Pharmacologic inhibition of KDM4 blocks expression of MYCN and the adrenergic CRC transcriptome with genome-wide induction of transcriptionally repressive H3K9me3, resulting in potent anticancer activity against neuroblastomas with MNA by inducing neuroblastic differentiation and apoptosis. Furthermore, a short-term KDM4 inhibition in combination with conventional, cytotoxic chemotherapy results in complete tumor responses of xenografts with MNA. Thus, KDM4 blockade may serve as a transformative strategy to target the adrenergic CRC dependencies in MNA neuroblastomas.

Metabolic reprogramming of cancer cells by JMJD6-mediated pre-mRNA splicing associated with therapeutic response to splicing inhibitor.

Dysregulated pre-mRNA splicing and metabolism are two hallmarks of MYC-driven cancers. Pharmacological inhibition of both processes has been extensively investigated as potential therapeutic avenues in preclinical and clinical studies. However, how pre-mRNA splicing and metabolism are orchestrated in response to oncogenic stress and therapies is poorly understood. Here, we demonstrate that jumonji domain containing 6, arginine demethylase, and lysine hydroxylase, JMJD6, acts as a hub connecting splicing and metabolism in MYC-driven human neuroblastoma. JMJD6 cooperates with MYC in cellular transformation of murine neural crest cells by physically interacting with RNA binding proteins involved in pre-mRNA splicing and protein homeostasis. Notably, JMJD6 controls the alternative splicing of two isoforms of glutaminase (GLS), namely kidney-type glutaminase (KGA) and glutaminase C (GAC), which are rate-limiting enzymes of glutaminolysis in the central carbon metabolism in neuroblastoma. Further, we show that JMJD6 is correlated with the anti-cancer activity of indisulam, a 'molecular glue' that degrades splicing factor RBM39, which complexes with JMJD6. The indisulam-mediated cancer cell killing is at least partly dependent on the glutamine-related metabolic pathway mediated by JMJD6. Our findings reveal a cancer-promoting metabolic program is associated with alternative pre-mRNA splicing through JMJD6, providing a rationale to target JMJD6 as a therapeutic avenue for treating MYC-driven cancers.

Sex and statin-related genetic associations at the PCSK9 gene locus: results of genome-wide association meta-analysis.

BACKGROUND: Proprotein convertase subtilisin/kexin type 9 (PCSK9) is a key player of lipid metabolism with higher plasma levels in women throughout their life. Statin treatment affects PCSK9 levels also showing evidence of sex-differential effects. It remains unclear whether these differences can be explained by genetics. METHODS: We performed genome-wide association meta-analyses (GWAS) of PCSK9 levels stratified for sex and statin treatment in six independent studies of Europeans (8936 women/11,080 men respectively 14,825 statin-free/5191 statin-treated individuals). Loci associated in one of the strata were tested for statin- and sex-interactions considering all independent signals per locus. Independent variants at the PCSK9 gene locus were then used in a stratified Mendelian Randomization analysis (cis-MR) of PCSK9 effects on low-density lipoprotein cholesterol (LDL-C) levels to detect differences of causal effects between the subgroups. RESULTS: We identified 11 loci associated with PCSK9 in at least one stratified subgroup (p < 1.0 × 10-6), including the PCSK9 gene locus and five other lipid loci: APOB, TM6SF2, FADS1/FADS2, JMJD1C, and HP/HPR. The interaction analysis revealed eight loci with sex- and/or statin-interactions. At the PCSK9 gene locus, there were four independent signals, one with a significant sex-interaction showing stronger effects in men (rs693668). Regarding statin treatment, there were two significant interactions in PCSK9 missense mutations: rs11591147 had stronger effects in statin-free individuals, and rs11583680 had stronger effects in statin-treated individuals. Besides replicating known loci, we detected two novel genome-wide significant associations: one for statin-treated individuals at 6q11.1 (within KHDRBS2) and one for males at 12q24.22 (near KSR2/NOS1), both with significant interactions. In the MR of PCSK9 on LDL-C, we observed significant causal estimates within all subgroups, but significantly stronger causal effects in statin-free subjects compared to statin-treated individuals. CONCLUSIONS: We performed the first double-stratified GWAS of PCSK9 levels and identified multiple biologically plausible loci with genetic interaction effects. Our results indicate that the observed sexual dimorphism of PCSK9 and its statin-related interactions have a genetic basis. Significant differences in the causal relationship between PCSK9 and LDL-C suggest sex-specific dosages of PCSK9 inhibitors.

The protein "proprotein convertase subtilisin/kexin type 9" (PCSK9) regulates the levels of low-density lipoprotein cholesterol (LDL-C) in blood, and thus, contributes to the risk of cardio-vascular diseases. Women tend to have higher PCSK9 plasma levels throughout their life, although the difference is smaller in patients under LDL-C lowering medication (e.g., statins). We investigated the interplay of genetics, statin-treatment and sex, using combined data from six European studies. We detected 11 genetic regions associated with PCSK9 levels, of which one was specific for women (at SLCO1B3, a statin-transporter gene), and three were specific for men (e.g., ALOX5, encoding a protein linked to chronic inflammatory diseases such as atherosclerosis). We also tested if statin use changed the genetic effect and found five genes only associated with PCSK9 levels in untreated participants. Variants in the gene encoding PCSK9 were most strongly associated and had heterogeneous effects in dependence on statin treatment and sex: On one hand, there were genetic variants with stronger effects in men than women. Those variants are also linked to sex-differential gene expression of PCSK9. On the other hand, there were also variants with treatment-depending effects, linked to protein structure and functionality of PCSK9. This indicates that the observed sexual and treatment-related effects on PCSK9 levels have a genetic basis. In addition, we compared the causal effects of PCSK9 on LDL-C levels between men and women and found a different response to statin treatment. This highlights the need for sex-sensitive dosages of lipid-lowering medication.

DUX4-induced HSATII transcription causes KDM2A/B-PRC1 nuclear foci and impairs DNA damage response.

Polycomb repressive complexes regulate developmental gene programs, promote DNA damage repair, and mediate pericentromeric satellite repeat repression. Expression of pericentromeric satellite repeats has been implicated in several cancers and diseases, including facioscapulohumeral dystrophy (FSHD). Here, we show that DUX4-mediated transcription of HSATII regions causes nuclear foci formation of KDM2A/B-PRC1 complexes, resulting in a global loss of PRC1-mediated monoubiquitination of histone H2A. Loss of PRC1-ubiquitin signaling severely impacts DNA damage response. Our data implicate DUX4-activation of HSATII and sequestration of KDM2A/B-PRC1 complexes as a mechanism of regulating epigenetic and DNA repair pathways.

Identification of epigenetically regulated genes involved in plant-virus interaction and their role in virus-triggered induced resistance.

BACKGROUND: Plant responses to a wide range of stresses are known to be regulated by epigenetic mechanisms. Pathogen-related investigations, particularly against RNA viruses, are however scarce. It has been demonstrated that Arabidopsis thaliana plants defective in some members of the RNA-directed DNA methylation (RdDM) or histone modification pathways presented differential susceptibility to the turnip mosaic virus. In order to identify genes directly targeted by the RdDM-related RNA Polymerase V (POLV) complex and the histone demethylase protein JUMONJI14 (JMJ14) during infection, the transcriptomes of infected mutant and control plants were obtained and integrated with available chromatin occupancy data for various epigenetic proteins and marks. RESULTS: A comprehensive list of virus-responsive gene candidates to be regulated by the two proteins was obtained. Twelve genes were selected for further characterization, confirming their dynamic regulation during the course of infection. Several epigenetic marks on their promoter sequences were found using in silico data, raising confidence that the identified genes are actually regulated by epigenetic mechanisms. The altered expression of six of these genes in mutants of the methyltransferase gene CURLY LEAF and the histone deacetylase gene HISTONE DEACETYLASE 19 suggests that some virus-responsive genes may be regulated by multiple coordinated epigenetic complexes. A temporally separated multiple plant virus infection experiment in which plants were transiently infected with one virus and then infected by a second one was designed to investigate the possible roles of the identified POLV- and JMJ14-regulated genes in wild-type (WT) plants. Plants that had previously been stimulated with viruses were found to be more resistant to subsequent virus challenge than control plants. Several POLV- and JMJ14-regulated genes were found to be regulated in virus induced resistance in WT plants, with some of them poisoned to be expressed in early infection stages. CONCLUSIONS: A set of confident candidate genes directly regulated by the POLV and JMJ14 proteins during virus infection was identified, with indications that some of them may be regulated by multiple epigenetic modules. A subset of these genes may also play a role in the tolerance of WT plants to repeated, intermittent virus infections.

KDM4A-AS1 Promotes Cell Proliferation, Migration, and Invasion via the miR-4306/STX6 Axis in Hepatocellular Carcinoma.

As a primary liver malignancy, hepatocellular carcinoma (HCC) is commonly induced by chronic liver disease and cirrhosis. Bioinformatics analysis reveals that long noncoding RNA KDM4A antisense RNA 1 (KDM4A-AS1) may be aberrantly expressed in HCC and its abnormal expression might influence prognosis in patients. We conducted this study to illustrate the functions and mechanism of KDM4A-AS1 in regulating HCC malignant cell behavior. KD-M4A-AS1, microRNA (miR)-4306 and messenger RNA syntaxin 6 (STX6) expression was examined by reverse transcription-quantitative polymerase chain reaction (RT-qPCR). HCC cell proliferation, apoptosis, migration, and invasion were measured by colony forming assays, flow cytometry, wound healing and Transwell assays. The interaction between genes was verified by RNA immunoprecipitation and luciferase reporter assays. Western blotting was performed to quantify protein expression of STX6 or apoptotic markers. KDM4A-AS1 was highly expressed in HCC cells and tissues. KDM4A-AS1 knockdown led to enhanced HCC cell apoptosis and suppressed HCC cell proliferation, migration, and invasion. MiR-4306 bound to and negatively regulated STX6. KDM4A-AS1 directly bound to miR-4306 and thus up-regulated STX6. STX6 overexpression reversed the inhibitory influence of KDM4A-AS1 depletion on HCC malignant behavior. KDM4A-AS1 promotes HCC cell migration, invasion, and growth by upregulating STX6 via miR-4306.

Kaempferol inhibits colorectal cancer metastasis through circ_0000345 mediated JMJD2C/ß-catenin signalling pathway.

BACKGROUND: Recurrence and metastasis are the main causes of disease deterioration in colorectal cancer (CRC) patients, yet efficient therapeutic strategies are lacking. Natural compounds for efficient antitumour therapeutics are becoming increasingly prominent. Kaempferol, one of the main components of flavonoids in plants, displays a variety of pharmacological activities. Our preliminary experiments suggested that kaempferol could inhibit CRC metastasis and is significantly associated with the ß-catenin signalling pathway. Moreover, we also defined the regulatory roles of JMJD2C in ß-catenin signalling in our previous work. PURPOSE: This study aims to reveal the mechanism by which kaempferol inhibits CRC progression and regulates the JMJD2C/ß-catenin signalling pathway. METHODS: The migratory capabilities of CRC cells after kaempferol intervention were measured by scratch wound healing and transwell assays. Circ_0000345 knockdown CRC stable cell lines were generated by lentivirus infection. The possible mechanism of kaempferol on circ_0000345 was verified by molecular-protein docking and verification program cellular thermal shift assay (CETSA). A dual luciferase reporter gene assay was carried out for the targeting relationship among circ_0000345, miR-205-5p and JMJD2C. Fluorescence in situ hybridization (FISH) was performed to determine the expression of circ_0000345 in tumour tissues. A pulmonary metastatic model of CRC in vitro was built to assess the antimetastatic effect and mechanism of kaempferol in vivo. RESULTS: In vitro, kaempferol inhibits the ability to migrate of CRC cells by reducing the activation of the JMJD2C/ß-catenin signalling pathway. MiR-205-5p is a key bridge for kaempferol to inhibit the expression of JMJD2C. The function of miR-205-5p is impeded by circ_0000345, which shows higher expression levels in human metastatic CRC tissues than nonmetastatic CRC tissues, and its formation is regulated by the RNA-binding proteins HNRNPK and HNRNPL. Mechanistically, kaempferol physically interacts with HNRNPK and HNRNPL to suppress JMJD2C by downregulating the expression of circ_0000345. In vivo, kaempferol suppresses CRC lung metastasis. Kaempferol inhibits the activation of JMJD2C/ß-catenin signalling through reducing the expression of circ_0000345 in the CRC lung metastasis model. CONCLUSION: Circ_0000345 enhances activation of the JMJD2C/ß-catenin signalling pathway through miR-205-5p to promote CRC metastasis. Kaempferol inhibits CRC metastasis through the circ_0000345-mediated JMJD2C/ß-catenin signalling pathway, and this effect is influenced as a direct consequence of the binding of kaempferol with HNRNPK and HNRNPL. This provides promising therapeutic and/or adjuvant agents for advanced CRC and sheds light on the multifaceted role of phytomedicine in cancer.

KDM5B promotes metastasis and epithelial-mesenchymal transition via Wnt/ß-catenin pathway in squamous cell carcinoma of the head and neck.

Metastasis determines clinical management decision and restricts the therapeutic efficiency in patients with squamous cell carcinoma of the head and neck (SCCHN). Epigenetic factor KDM5B serves as an oncogene in multiple cancers. However, its role in SCCHN metastasis remains unclear. Our previous study showed that KDM5B is significantly elevated in SCCHN tissue and is positively correlated with metastasis and recurrence. KDM5B overexpression predicted a poor prognosis in both disease-free survival and overall survival, which served as an independent prognostic factor in SCCHN patients. This study further investigates the exact impact of KDM5B in metastasis of SCCHN. We found that KDM5B knockdown significantly inhibits the migration and invasion of SCCHN cells both in vitro and in vivo. On the contrary, forced expression of KDM5B leads to enhanced migration and invasion, accompanied by canonical alterations of epithelial-mesenchymal transition (EMT). Mechanism investigations demonstrated that KDM5B activates Wnt/ß-catenin pathway, and inhibition of Wnt/ß-catenin pathway via a small molecule inhibitor iCRT-14 partially reverses the enhanced migratory and invasive ability caused by KDM5B in SCCHN cells. Together, our data indicate that KDM5B promotes EMT and metastasis via Wnt/ß-catenin pathway in SCCHN, suggesting that KDM5B may be a potential therapeutic target and prognosis biomarker in SCCHN.

Sex-Hormone-Binding Globulin Gene Polymorphisms and Breast Cancer Risk in Caucasian Women of Russia.

In our work, the associations of GWAS (genome-wide associative studies) impact for sex-hormone-binding globulin (SHBG)-level SNPs with the risk of breast cancer (BC) in the cohort of Caucasian women of Russia were assessed. The work was performed on a sample of 1498 women (358 BC patients and 1140 control (non BC) subjects). SHBG correlated in previously GWAS nine polymorphisms such as rs780093 GCKR, rs17496332 PRMT6, rs3779195 BAIAP2L1, rs10454142 PPP1R21, rs7910927 JMJD1C, rs4149056 SLCO1B1, rs440837 ZBTB10, rs12150660 SHBG, and rs8023580 NR2F2 have been genotyped. BC risk effects of allelic and non-allelic SHBG-linked gene SNPs interactions were detected by regression analysis. The risk genetic factor for BC developing is an SHBG-lowering allele variant C rs10454142 PPP1R21 ([additive genetic model] OR = 1.31; 95%CI = 1.08-1.65; pperm = 0.024; power = 85.26%), which determines 0.32% of the cancer variance. Eight of the nine studied SHBG-related SNPs have been involved in cancer susceptibility as part of nine different non-allelic gene interaction models, the greatest contribution to which is made by rs10454142 PPP1R21 (included in all nine models, 100%) and four more SNPs-rs7910927 JMJD1C (five models, 55.56%), rs17496332 PRMT6 (four models, 44.44%), rs780093 GCKR (four models, 44.44%), and rs440837 ZBTB10 (four models, 44.44%). For SHBG-related loci, pronounced functionality in the organism (including breast, liver, fibroblasts, etc.) was predicted in silico, having a direct relationship through many pathways with cancer pathophysiology. In conclusion, our results demonstrated the involvement of SHBG-correlated genes polymorphisms in BC risk in Caucasian women in Russia.

Targeting KDM4 family epigenetically triggers antitumour immunity via enhancing tumour-intrinsic innate sensing and immunogenicity.

Despite the remarkable clinical efficacy of cancer immunotherapy, considerable patients fail to benefit from it due to primary or acquired resistance. Tumours frequently hijack diverse epigenetic mechanisms to evade immune detection, thereby highlighting the potential for pharmacologically targeting epigenetic regulators to restore the impaired immunosurveillance and re-sensitise tumours to immunotherapy. Herein, we demonstrated that KDM4-targeting chemotherapeutic drug JIB-04, epigenetically triggered the tumour-intrinsic innate immune responses and immunogenic cell death (ICD), resulting in impressive antitumour effects. Specifically, JIB-04 induced H3K9 hypermethylation through specific inhibition of the KDM4 family (KDM4A-D), leading to impaired DNA repair signalling and subsequent DNA damage. As a result, JIB-04 not only activated the tumour-intrinsic cyclic GMP-AMP synthase (cGAS)-STING pathway via DNA-damage-induced cytosolic DNA accumulation, but also promoted ICD, releasing numerous damage-associated molecular patterns. Furthermore, JIB-04 induced adaptive resistance through the upregulation of programmed death-ligand 1 (PD-L1), which could be overcome with additional PD-L1 blockade. In human tumours, KDM4B expression was negatively correlated with clinical outcomes, type I interferon signatures, and responses to immunotherapy. In conclusion, our results demonstrate that targeting KDM4 family can activate tumour-intrinsic innate sensing and immunogenicity, and synergise with immunotherapy to improve antitumour outcomes.