ERK3 is involved in regulating cardiac fibroblast function.

ERK3/MAPK6 activates MAP kinase-activated protein kinase (MK)-5 in selected cell types. Male MK5 haplodeficient mice show reduced hypertrophy and attenuated increase in Col1a1 mRNA in response to increased cardiac afterload. In addition, MK5 deficiency impairs cardiac fibroblast function. This study determined the effect of reduced ERK3 on cardiac hypertrophy following transverse aortic constriction (TAC) and fibroblast biology in male mice. Three weeks post-surgery, ERK3, but not ERK4 or p38α, co-immunoprecipitated with MK5 from both sham and TAC heart lysates. The increase in left ventricular mass and myocyte diameter was lower in TAC-ERK3+/- than TAC-ERK3+/+ hearts, whereas ERK3 haploinsufficiency did not alter systolic or diastolic function. Furthermore, the TAC-induced increase in Col1a1 mRNA abundance was diminished in ERK3+/- hearts. ERK3 immunoreactivity was detected in atrial and ventricular fibroblasts but not myocytes. In both quiescent fibroblasts and "activated" myofibroblasts isolated from adult mouse heart, siRNA-mediated knockdown of ERK3 reduced the TGF-ß-induced increase in Col1a1 mRNA. In addition, intracellular type 1 collagen immunoreactivity was reduced following ERK3 depletion in quiescent fibroblasts but not myofibroblasts. Finally, knocking down ERK3 impaired motility in both atrial and ventricular myofibroblasts. These results suggest that ERK3 plays an important role in multiple aspects of cardiac fibroblast biology.

Long non-coding RNA DANCR increases spinal cord neuron apoptosis and inflammation of spinal cord injury by mediating the microRNA-146a-5p/MAPK6 axis.

OBJECTIVE: This research was to unravel the impact of the lncRNA differentiation antagonizing non-protein coding RNA (DANCR)/microRNA (miR)-146a-5p/mitogen-activated protein kinase 6 (MAPK6) axis on spinal cord injury (SCI). METHODS: SCI mouse models were established and injected with si-DANCR or miR-146a-5p agomir. The recovery of motor function was assessed by Basso Mouse Scale. SCI was pathologically evaluated, and serum inflammatory factors were measured in SCI mice. Mouse spinal cord neurons were injured by H2O2 and transfected, followed by assessment of proliferation and apoptosis. DANCR, miR-146a-5p, and MAPK6 in tissues and cells were detected, as well as their relationship. RESULTS: DANCR increased and miR-146a-5p decreased in SCI. Silencing DANCR or enhancing miR-146a-5p stimulated the proliferation of mouse spinal cord neurons and reduced apoptosis. DANCR was bound to miR-146a-5p to target MAPK6. DANCR affected the proliferation and apoptosis of spinal cord neurons by mediating the miR-146a-5p/MAPK6 axis. Downregulating DANCR or upregulating miR-146a-5p improved inflammation, the destruction of spinal cord tissue structure, and apoptosis in SCI mice. CONCLUSION: DANCR affects spinal cord neuron apoptosis and inflammation of SCI by mediating the miR-146a-5p/MAPK6 axis.

Exosomes from miR-374a-5p-modified mesenchymal stem cells inhibit the progression of renal fibrosis by regulating MAPK6/MK5/YAP axis.

Chronic kidney disease (CKD) in clinical is defined as a gradual loss of kidney function for more than 3 months. The pathologic course of CKD is characterized by extensive renal fibrosis; thus, preventing renal fibrosis is vital for the treatment of CKD. It has been reported that microRNA (miR)-374a-5p was under-expressed in renal venous blood samples from patients with CKD. In addition, it exhibited anti-apoptotic effects in renal tissues suggesting that miR-374a-5p may play an important role in CKD. However, it is not clear whether miR-374a-5p could be delivered to renal cells by exosomes and exerts anti-renal fibrosis effects. To mimic renal fibrosis in vitro, human renal tubular epithelial cell lines (HK-2 cells) were treated by transforming growth factor-ß (TGF-ß) 1. Reverse transcription-quantitative polymerase-chain reaction (RT-qPCR) or Western blot was carried out to evaluate the mechanism by which miR-374a-5p regulated the development of renal fibrosis. Next, exosomes were isolated using with ultracentrifugation method, and the relationship between miR-374a-5p and MAPK6 was evaluated using dual-Luciferase a reporter assay system. The results indicated TGF-ß1 significantly down-regulated the expression of miR-374a-5p in HK-2 cells and miR-374a-5p agomir remarkably inhibited the progression of fibrosis in vitro. In addition, exosomal miR-374a-5p could be internalized by HK-2 cells and obviously enhanced the level of miR-374a-5p in HK-2 cells. Furthermore, exosomal miR-374a-5p prevented the progression of renal fibrosis in vivo by regulating MAPK6/MK5/YAP axis. In conclusion, exosomal miR-374a-5p inhibited the progression of renal fibrosis by regulating MAPK6/MK5/YAP axis.

Conditional ERK3 overexpression cooperates with PTEN deletion to promote lung adenocarcinoma formation in mice.

ERK3, officially known as mitogen-activated protein kinase 6 (MAPK6), is a poorly studied mitogen-activated protein kinase (MAPK). Recent studies have revealed the upregulation of ERK3 expression in cancer and suggest an important role for ERK3 in promoting cancer cell growth and invasion in some cancers, in particular lung cancer. However, it is unknown whether ERK3 plays a role in spontaneous tumorigenesis in vivo. To determine the role of ERK3 in lung tumorigenesis, we created a conditional ERK3 transgenic mouse line in which ERK3 transgene expression is controlled by Cre recombinase. By crossing these transgenic mice with a mouse line harboring a lung tissue-specific Cre recombinase transgene driven by a club cell secretory protein gene promoter (CCSP-iCre), we have found that conditional ERK3 overexpression cooperates with phosphatase and tensin homolog (PTEN) deletion to induce the formation of lung adenocarcinomas (LUADs). Mechanistically, ERK3 overexpression stimulates activating phosphorylations of erb-b2 receptor tyrosine kinases 2 and 3 (ERBB2 and ERBB3) by upregulating Sp1 transcription factor (SP1)-mediated gene transcription of neuregulin 1 (NRG1), a potent ligand for ERBB2/ERBB3. Our study has revealed a bona fide tumor-promoting role for ERK3 using genetically engineered mouse models. Together with previous findings showing the roles of ERK3 in cultured cells and in a xenograft lung tumor model, our findings corroborate that ERK3 acts as an oncoprotein in promoting LUAD development and progression.

Endurance exercise training-responsive miR-19b-3p improves skeletal muscle glucose metabolism.

Skeletal muscle is a highly adaptable tissue and remodels in response to exercise training. Using short RNA sequencing, we determine the miRNA profile of skeletal muscle from healthy male volunteers before and after a 14-day aerobic exercise training regime. Among the exercise training-responsive miRNAs identified, miR-19b-3p was selected for further validation. Overexpression of miR-19b-3p in human skeletal muscle cells increases insulin signaling, glucose uptake, and maximal oxygen consumption, recapitulating the adaptive response to aerobic exercise training. Overexpression of miR-19b-3p in mouse flexor digitorum brevis muscle enhances contraction-induced glucose uptake, indicating that miR-19b-3p exerts control on exercise training-induced adaptations in skeletal muscle. Potential targets of miR-19b-3p that are reduced after aerobic exercise training include KIF13A, MAPK6, RNF11, and VPS37A. Amongst these, RNF11 silencing potentiates glucose uptake in human skeletal muscle cells. Collectively, we identify miR-19b-3p as an aerobic exercise training-induced miRNA that regulates skeletal muscle glucose metabolism.

Bacterial leaf streak 1 encoding a mitogen-activated protein kinase confers resistance to bacterial leaf streak in rice.

Bacterial leaf streak (BLS) is a major bacterial disease of rice. Utilization of host genetic resistance has become one of the most important strategies for controlling BLS. However, only a few resistance genes have been characterized. Previously, a recessive BLS resistance gene bls1 was roughly mapped on chromosome 6. Here, we further delineated bls1 to a 21 kb region spanning four genes. Genetic analysis confirmed that the gene encoding a mitogen-activated protein kinase (OsMAPK6) is the target of the allelic genes BLS1 and bls1. Overexpression of BLS1 weakened resistance to the specific Xanthomonas oryzae pv. oryzicola (Xoc) strain JZ-8, while low expression of bls1 increased resistance. However, both overexpression of BLS1 and low expression of bls1 could increase no-race-specific broad-spectrum resistance. These results indicate that BLS1 and bls1 negatively regulate race-specific resistance to Xoc strain JZ-8 but positively and negatively control broad-spectrum resistance, respectively. Subcellular localization demonstrated that OsMAPK6 was localized in the nucleus. RGA4, which is known to mediate resistance to Xoc, is the potential target of OsMAPK6. Overexpression of BLS1 and low expression of bls1 showed increase in salicylic acid and induced expression of defense-related genes, simultaneously increasing broad-spectrum resistance. Moreover, low expression of bls1 showed increase an in jasmonic acid and abscisic acid, in company with an increase in resistance to Xoc strain JZ-8. Collectively, our study provides new insights into the understanding of BLS resistance and facilitates the development of rice host-resistant cultivars.

ERK3 is transcriptionally upregulated by ∆Np63α and mediates the role of ∆Np63α in suppressing cell migration in non-melanoma skin cancers.

BACKGROUND: p63, a member of the p53 gene family, is an important regulator for epithelial tissue growth and development. ∆Np63α is the main isoform of p63 and highly expressed in Non-melanoma skin cancer (NMSC). Extracellular signal-regulated kinase 3 (ERK3) is an atypical mitogen-activated protein kinase (MAPK) whose biochemical features and cellular regulation are distinct from those of conventional MAPKs such as ERK1/2. While ERK3 has been shown to be upregulated in lung cancers and head and neck cancers, in which it promotes cancer cell migration and invasion, little is known about the implication of ERK3 in NMSCs. METHODS: Fluorescent immunohistochemistry was performed to evaluate the expression levels of ΔNp63α and ERK3 in normal and NMSC specimens. Dunnett's test was performed to compare mean fluorescence intensity (MFI, indicator of expression levels) of p63 or ERK3 between normal cutaneous samples and NMSC samples. A mixed effects (ANOVA) test was used to determine the correlation between ΔNp63α and ERK3 expression levels (MFI). The regulation of ERK3 by ΔNp63α was studied by qRT-PCR, Western blot and luciferase assay. The effect of ERK3 regulation by ΔNp63α on cell migration was measured by performing trans-well migration assay. RESULTS: The expression level of ∆Np63α is upregulated in NMSCs compared to normal tissue. ERK3 level is significantly upregulated in AK and SCC in comparison to normal tissue and there is a strong positive correlation between ∆Np63α and ERK3 expression in normal skin and skin specimens of patients with AK, SCC or BCC. Further, we found that ∆Np63α positively regulates ERK3 transcript and protein levels in A431 and HaCaT skin cells, underlying the upregulation of ERK3 expression and its positive correlation with ∆Np63α in NMSCs. Moreover, similar to the effect of ∆Np63α depletion, silencing ERK3 greatly enhanced A431 cell migration. Restoration of ERK3 expression under the condition of silencing ∆Np63α counteracted the increase in cell migration induced by the depletion of ∆Np63α. Mechanistically, ERK3 inhibits the phosphorylation of Rac1 G-protein and the formation of filopodia of A431 skin SCC cells. CONCLUSIONS: ERK3 is positively regulated by ∆Np63α and mediates the role of ∆Np63α in suppressing cell migration in NMSC.

miR­653­5p suppresses the growth and migration of breast cancer cells by targeting MAPK6.

Breast cancer is the worldwide leading cause of cancer­related deaths among women. Increasing evidence has demonstrated that microRNAs (miRNAs) play critical roles in the carcinogenesis and progression of breast cancer. miR­653­5p was previously reported to be involved in cell proliferation and apoptosis. However, the role of miR­653­5p in the progression of breast cancer has not been studied. In the present study, it was found that overexpression of miR­653­5p significantly inhibited the proliferation, migration and invasion of breast cancer cells in vitro. Moreover, overexpression of miR­653­5p promoted cell apoptosis in breast cancer by regulating the Bcl­2/Bax axis and caspase­9 activation. Additionally, the epithelial­mesenchymal transition and activation of the Akt/mammalian target of rapamycin signaling pathway were also inhibited by miR­653­5p. Furthermore, the data demonstrated that miR­653­5p directly targeted mitogen­activated protein kinase 6 (MAPK6) and negatively regulated its expression in breast cancer cells. Upregulation of MAPK6 could overcome the inhibitory effects of miR­653­5p on cell proliferation and migration in breast cancer. In conclusion, this study suggested that miR­653­5p functions as a tumor suppressor by targeting MAPK6 in the progression of breast cancer, and it may be a potential target for breast cancer therapy.

LncRNA GAS6-AS2 promotes non-small-cell lung cancer cell proliferation via regulating miR-144-3p/ MAPK6 axis.

The function of a new long non-coding RNA GAS6-AS2 in non-small cell lung cancer (NSCLC) is not fully understood. In this study, GAS6-AS2 was identified, and its roles as well as mechanisms in regulating proliferation of NSCLCs cells were investigated. qRT-PCR was used to analyze GAS6-AS2, miR-144-3p, and MAPK6 expression. Protein expression was detected by Western blotting. Cell Counting Kit-8 (CCK8) assay was used to examine the cell proliferation ability. The interaction between GAS6-AS2 and miR-144-3p was confirmed by dual-luciferase reporter assay and RNA pull down assay. A xenograft model was constructed to monitor the mice NSCLC tumor growth in vivo. GAS6-AS2 was up-regulated, while miR-144-3p was suppressed in NSCLC cells compared with normal lung cells. GAS6-AS2 suppression could inhibit the progression of NSCLC cells, and miR-144-3p could attenuate the effect. GAS6-AS2 could function as a competitive endogenous RNA (ceRNA) via direct sponging miR-144-3p-3p, which further regulating the expression of MAPK6. The knockdown of GAS6-AS2 could greatly suppress the tumor growth of NSCLC in vivo. GAS6-AS2 up-regulated MAPK6 by sponging miR-144-3p in NSCLC tissues and cells. Thus, GAS6-AS2 is an effective therapeutic target in NSCLC.

let7f­5p attenuates inflammatory injury in in vitro pneumonia models by targeting MAPK6.

Pneumonia accounts for ~1.3 million mortalities in children per year worldwide. MicroRNAs are implicated in several diseases, including cancer and pneumonia; however, the role of let7f­5p in pneumonia is not completely understood. In the present study, lipopolysaccharide (LPS) was used to establish an in vitro pneumonia model in A549 and WI­38 cells. The reverse transcription­quantitative PCR (RT­qPCR) and western blotting results demonstrated that let7f­5p expression levels were significantly decreased, whereas MAPK6 expression levels were significantly increased in the peripheral venous blood of patients with pneumonia and in LPS­induced A549 and WI­38 cells compared with healthy volunteers and control cells, respectively. Furthermore, the dual­luciferase reporter assay demonstrated that let7f­5p targeted the 3'­untranslated region of MAPK6. The ELISA and RT­qPCR results demonstrated that let7f­5p mimic ameliorated LPS­induced inflammatory injury in A549 and WI­38 cells, as demonstrated by decreased expression levels of proinflammatory cytokines, including TNF­α and IL­6. In addition, the Cell Counting Kit­8 assay results indicated that let7f­5p mimic ameliorated LPS­induced reductions in cell viability, and the western blotting results demonstrated that let7f­5p mimic reversed LPS­induced activation of the STAT3 signaling pathway. Notably, the aforementioned let7f­5p­mediated effects were reversed by MAPK6 overexpression. Collectively, the results of the present study suggested that let7f­5p inhibited inflammation by targeting MAPK6 in the in vitro pneumonia model, thus let7f­5p may serve as a potential novel therapeutic target for pneumonia.

ERK3/MAPK6 is required for KRAS-mediated NSCLC tumorigenesis.

KRAS is one of the most frequently mutated oncogenes, especially in lung cancers. Targeting of KRAS directly or the downstream effector signaling machinery is of prime interest in treating lung cancers. Here, we uncover that ERK3, a ubiquitously expressed atypical MAPK, is required for KRAS-mediated NSCLC tumors. ERK3 is highly expressed in lung cancers, and oncogenic KRAS led to the activation and stabilization of the ERK3 protein. In particular, phosphorylation of serine 189 in the activation motif of ERK3 is significantly increased in lung adenocarcinomas in comparison to adjacent normal controls in patients. Loss of ERK3 prevents the anchorage-independent growth of KRAS G12C-transformed human bronchial epithelial cells. We further find that loss of ERK3 reduces the oncogenic growth of KRAS G12C-driven NSCLC tumors in vivo and that the kinase activity of ERK3 is required for KRAS-driven oncogenesis in vitro. Our results demonstrate an obligatory role for ERK3 in NSCLC tumor progression and suggest that ERK3 kinase inhibitors can be pursued for treating KRAS G12C-driven tumors.

CircRNA_100395 protects breast carcinoma deterioration by targeting MAPK6.

OBJECTIVE: This study aims to uncover the differential expression of circRNA_100395 in breast carcinoma specimens, and its regulatory effect on cancer cell phenotypes. The role of circRNA_100395 in affecting breast carcinoma progression and the molecular mechanism are explored as well. PATIENTS AND METHODS: CircRNA_100395 expressions in breast carcinoma and paracancerous tissues were detected. The influence of circRNA_100395 level on clinical indicators of breast carcinoma patients was analyzed. In vitro regulations of circRNA_100395 on phenotypes of breast carcinoma cells were examined by CCK-8, colony formation, and transwell assay. The interaction between circRNA_100395 and MAPK6 was confirmed by Dual-Luciferase reporter assay and rescue assays. RESULTS: CircRNA_100395 was downregulated in breast carcinoma tissues and cell lines. Its level was negatively correlated to tumor staging and tumor size of breast carcinoma. Overexpression of circRNA_100395 in SKBR3 and MDA-MB-231 cells weakened proliferative and migratory abilities. MAPK6 was the target gene of circRNA_100395. Overexpression of MAPK6 reversed the anti-cancer effect of circRNA_100395 on breast carcinoma. CONCLUSIONS: CircRNA_100395 serves as an anti-cancer gene in breast carcinoma progression by targeting MAPK6, and its level is negatively correlated to tumor staging and tumor size of breast carcinoma. CircRNA_100395 can be utilized as a potential biomarker and therapeutic target of breast carcinoma.

The C-Terminus Tail Regulates ERK3 Kinase Activity and Its Ability in Promoting Cancer Cell Migration and Invasion.

Extracellular signal-regulated kinase 3 (ERK3) is an atypical member of the mitogen-activated protein kinase (MAPK) family. It harbors a kinase domain in the N-terminus and a long C-terminus extension. The C-terminus extension comprises a conserved in ERK3 and ERK4 (C34) region and a unique C-terminus tail, which was shown to be required for the interaction of ERK3 with the cytoskeletal protein septin 7. Recent studies have elucidated the role of ERK3 signaling in promoting the motility and invasiveness of cancer cells. However, little is known about the intramolecular regulation of the enzymatic activity and cellular functions of ERK3. In this study, we investigated the role of the elongated C-terminus extension in regulating ERK3 kinase activity and its ability to promote cancer cell migration and invasion. Our study revealed that the deletion of the C-terminus tail greatly diminishes the ability of ERK3 to promote the migration and invasion of lung cancer cells. We identified two molecular mechanisms underlying this effect. Firstly, the deletion of the C-terminus tail decreases the kinase activity of ERK3 towards substrates, including the oncogenic protein steroid receptor co-activator 3 (SRC-3), an important downstream target for ERK3 signaling in cancer. Secondly, in line with the previous finding that the C-terminus tail mediates the interaction of ERK3 with septin 7, we found that the depletion of septin 7 abolished the ability of ERK3 to promote migration, indicating that septin 7 acts as a downstream effector for ERK3-induced cancer cell migration. Taken together, the findings of this study advance our understanding of the molecular regulation of ERK3 signaling by unraveling the role of the C-terminus tail in regulating ERK3 kinase activity and functions in cancer cells. These findings provide useful insights for the development of therapeutic agents targeting ERK3 signaling in cancer.

LncRNA SNHG1 regulates vascular endothelial cell proliferation and angiogenesis via miR-196a.

Inflammatory cytokines are important protagonists in the formation of atherosclerotic plaques, triggering effects throughout the atherosclerotic vessels due to the destruction in proliferation, migration and angiogenesis of endothelial cells. In this study, we found SNHG1 is upregulated in TNF-α-treated HUVECs. We silenced SNHG1 and found it inhibited vascular endothelial cell proliferation and angiogenesis. In the other hand, exogenetic overexpression of SNHG1 promotes proliferation, migration and angiogenesis. Then we demonstrated that SNHG1 may interact directly with miR-196a to act as a miR-196a sponge. Further, MAPK6 were predicted to be the target of miR-196a. So we blocked miR-196a, which increased expression level of MAPK6, enhanced cell proliferation, migration and angiogenesis. These data indicated that SNHG1/miR-196a/MAPK6 axis may take a part in autophagy regulation in TNF-α-treated HUVECs. The subsequent rescue experiments come to the results ascertained the specificity of SNHG1/miR-196a/MAPK6 axis in regulating MAPK6. Overall, our findings demonstrate a novel mechanism by which SNHG1 overexpression protects the function of HUVECs, which may delay the progression of AS. SNHG1/miR-196a/MAPK6 axis may be of therapeutic significance in AS.

Circ_0000285 promotes podocyte injury through sponging miR-654-3p and activating MAPK6 in diabetic nephropathy.

Recently, increasing evidence has reported that circRNAs are non-coding RNAs and they bind with the corresponding miRNAs to modulate the target genes. However, the detailed role of circRNAs in the pathogenesis of DN still remains poorly known. Currently, we aimed to study how circ_0000285 functions in DN development. We found that circ_0000285 was significantly increased in DN mice models and mouse podocytes incubated with HG. Then, circ_0000285 was overexpressed in mouse podocytes and we observed that overexpression of circ_0000285 promoted podocytes injury. Moreover, miR-654-3p was precited as a target of circ_0000285. It was shown that circ_0000285 was strongly pulled down by circ_0000285 specific probe and circ_0000285 specific probe was used to successfully enrich miR-654-3p. In addition, we reported that miR-654-3p was obviously down-regulated in DN. Inhibitors of miR-654-3p greatly reversed the effects of circ_0000285 siRNA on podocytes injury. Moreover, the inflammation release was restrained by loss of circ_0000285, while induced by miR-654-3p inhibitors. IL-6, L-1ß and TNF-α level was remarkably depressed by the knockdown of circ_0000285 and miR-654-3p inhibitors induced that. Furthermore, MAPK6 was confirmed as a direct downstream target of miR-654-3p. As shown, MAPK6 was markedly suppressed by circ_0000285 siRNA, which was rescued by the decrease of miR-654-3p. These findings revealed that circ_0000285 promoted podocyte injury via sponging miR-654-3p and activating MAPK6 in DN.

The adrenergic-induced ERK3 pathway drives lipolysis and suppresses energy dissipation.

Obesity-induced diabetes affects >400 million people worldwide. Uncontrolled lipolysis (free fatty acid release from adipocytes) can contribute to diabetes and obesity. To identify future therapeutic avenues targeting this pathway, we performed a high-throughput screen and identified the extracellular-regulated kinase 3 (ERK3) as a hit. We demonstrated that ß-adrenergic stimulation stabilizes ERK3, leading to the formation of a complex with the cofactor MAP kinase-activated protein kinase 5 (MK5), thereby driving lipolysis. Mechanistically, we identified a downstream target of the ERK3/MK5 pathway, the transcription factor FOXO1, which promotes the expression of the major lipolytic enzyme ATGL. Finally, we provide evidence that targeted deletion of ERK3 in mouse adipocytes inhibits lipolysis, but elevates energy dissipation, promoting lean phenotype and ameliorating diabetes. Thus, ERK3/MK5 represents a previously unrecognized signaling axis in adipose tissue and an attractive target for future therapies aiming to combat obesity-induced diabetes.

Erk1/2 inactivation promotes a rapid redistribution of COP1 and degradation of COP1 substrates.

Anthrax lethal toxin (LT) is a protease virulence factor produced by Bacillus anthracis that is required for its pathogenicity. LT treatment causes a rapid degradation of c-Jun protein that follows inactivation of the MEK1/2-Erk1/2 signaling pathway. Here we identify COP1 as the ubiquitin E3 ligase that is essential for LT-induced c-Jun degradation. COP1 knockdown using siRNA prevents degradation of c-Jun, ETV4, and ETV5 in cells treated with either LT or the MEK1/2 inhibitor, U0126. Immunofluorescence staining reveals that COP1 preferentially localizes to the nuclear envelope, but it is released from the nuclear envelope into the nucleoplasm following Erk1/2 inactivation. At baseline, COP1 attaches to the nuclear envelope via interaction with translocated promoter region (TPR), a component of the nuclear pore complex. Disruption of this COP1-TPR interaction, through Erk1/2 inactivation or TPR knockdown, leads to rapid COP1 release from the nuclear envelope into the nucleoplasm where it degrades COP1 substrates. COP1-mediated degradation of c-Jun protein, combined with LT-mediated blockade of the JNK1/2 signaling pathway, inhibits cellular proliferation. This effect on proliferation is reversed by COP1 knockdown and ectopic expression of an LT-resistant MKK7-4 fusion protein. Taken together, this study reveals that the nuclear envelope acts as a reservoir, maintaining COP1 poised for action. Upon Erk1/2 inactivation, COP1 is rapidly released from the nuclear envelope, promoting the degradation of its nuclear substrates, including c-Jun, a critical transcription factor that promotes cellular proliferation. This regulation allows mammalian cells to respond rapidly to changes in extracellular cues and mediates pathogenic mechanisms in disease states.

Expression and Secretion of an Atrial Natriuretic Peptide in Beige-Like 3T3-L1 Adipocytes.

The browning of white adipose tissue (beige adipocytes) stimulates energy expenditure. Omega-3 fatty acids have been shown to induce thermogenic action in adipocytes via G-protein coupled receptor 120 (GPR120). Atrial natriuretic peptide (ANP) is a peptide hormone that plays the role of maintaining normal blood pressure in kidneys to inhibit Na+ reuptake. Recently, ANP was found to induce adipocyte browning by binding to NPR1, an ANP receptor. However, the expression of ANP in adipocytes has not yet been studied. Therefore, in this study, we investigate the expression of ANP in beige-like adipocytes induced by docosahexaenoic acids (DHA), T3, or a PPAR agonist, rosiglitazone. First, we found that brown adipocyte-specific genes were upregulated in beige-like adipocytes. DHA promoted ANP expression in beige-like cells, whereas DHA-induced ANP expression was abolished by GPR120 knockout. ANP secretion of beige-like adipocytes was increased via PKC/ERK1/2 signaling in the GPR120 pathway. Furthermore, ANP secreted from beige-like adipocytes acted on HEK-293 cells, the recipient cells, leading to increased cGMP activity. After the NPR1 knockdown of HEK-293 cells, cGMP activity was not changed. Taken together, our findings indicate that beige-like adipocytes induce ANP secretion, which may contribute to improving obesity-associated metabolic disease.

The receptor-receptor interaction between mGluR1 receptor and NMDA receptor: a potential therapeutic target for protection against ischemic stroke.

Ischemic stroke is one of the leading causes of long-term disability worldwide. It arises when the blood flow to the brain is severely impaired, causing brain infarction. The current therapies for ischemic stroke are tissue plasminogen activator and mechanical thrombectomy, which re-establishes blood circulation to the brain but offers no neuroprotective effects. Excitotoxicity, particularly through the N-methyl-d-aspartate receptor (NMDAR), has been heavily implicated in the pathophysiology of brain infarction resulting from ischemic stroke. Here we investigated the interaction between NMDAR and metabotropic glutamate receptor 1 (mGluR1) as a novel target to develop potential neuroprotective agents for ischemic stroke. Through coimmunoprecipitation and affinity binding assay, we revealed that the interaction is mediated through 2 distinct sites on the mGluR1 C terminus. We then found that the disruption of mGluR1-GluN2A subunit of NMDAR (GluN2A) protected the primary mouse hippocampal neurons against NMDAR-mediated excitotoxicity and reversed the NMDAR-mediated regulation of ERK1/2 in rat hippocampal slices. The same protection was also observed in an animal model of ischemic stroke, alleviating brain infarction and yielding better motor recovery. These findings confirmed the existence of a receptor-receptor interaction between NMDAR and mGluR1, implicating this interconnection as a potential treatment target site for ischemic stroke.-Lai, T. K. Y., Zhai, D. Su, P., Jiang, A., Boychuk, J., Liu, F. The receptor-receptor interaction between mGluR1 receptor and NMDA receptor: a potential therapeutic target for protection against ischemic stroke.

MicroRNA-374a protects against myocardial ischemia-reperfusion injury in mice by targeting the MAPK6 pathway.

AIMS: Clinical treatment strategies for patients with myocardial ischemia typically include coronary artery recanalization to restore myocardial blood supply. However, myocardial reperfusion insult often induces oxidative stress and inflammation, which further leads to apoptosis and necrosis of myocardial cells. Increasing evidence suggests that microRNAs (miRNAs) participate in the pathological and physiological processes associated with myocardial ischemia reperfusion. MAIN METHODS: In this study, we established a myocardial H/R H9C2 cell model and a mouse I/R model to detect molecules implicated in myocardial I/R regulation and to determine the underlying signal transduction pathways. KEY FINDINGS: Herein, we showed that the expression of miR-374a-5p decreased in a myocardial cell model (H9C2 cells) of hypoxia/reoxygenation (H/R) and mouse model of ischemia/reperfusion (I/R). Alternatively, overexpression of miR-374a-5p was found to ameliorate myocardial cell damage within both in vivo and in vitro models of ischemia. Further, mitogen-activated protein kinase 6 (MAPK6) was identified as a direct target of miR-374a-5p. Thus, by targeting MAPK6, miR-374a-5p was found to negatively regulate MAPK6 expression. However, up-regulation of MAPK6 functioned to inhibit the previously observed protective effect of miR-374a-5p in the H9C2 H/R model. SIGNIFICANCE: Taken together, our study suggests that miR-374a-5p may have protective effects against cardiac I/R injury in vivo, and H/R injury in vitro, thereby providing novel insights into the molecular mechanisms associated with ischemia/reperfusion injury and a potential novel therapeutic target.