ARAF protein kinase activates RAS by antagonizing its binding to RASGAP NF1.

RAF protein kinases are effectors of the GTP-bound form of small guanosine triphosphatase RAS and function by phosphorylating MEK. We showed here that the expression of ARAF activated RAS in a kinase-independent manner. Binding of ARAF to RAS displaced the GTPase-activating protein NF1 and antagonized NF1-mediated inhibition of RAS. This reduced ERK-dependent inhibition of RAS and increased RAS-GTP. By this mechanism, ARAF regulated the duration and consequences of RTK-induced RAS activation and supported the RAS output of RTK-dependent tumor cells. In human lung cancers with EGFR mutation, amplification of ARAF was associated with acquired resistance to EGFR inhibitors, which was overcome by combining EGFR inhibitors with an inhibitor of the protein tyrosine phosphatase SHP2 to enhance inhibition of nucleotide exchange and RAS activation.

Dynamics of an incoherent feedforward loop drive ERK-dependent pattern formation in the early Drosophila embryo.

Positional information in development often manifests as stripes of gene expression, but how stripes form remains incompletely understood. Here, we use optogenetics and live-cell biosensors to investigate the posterior brachyenteron (byn) stripe in early Drosophila embryos. This stripe depends on interpretation of an upstream ERK activity gradient and the expression of two target genes, tailless (tll) and huckebein (hkb), that exert antagonistic control over byn. We find that high or low doses of ERK signaling produce transient or sustained byn expression, respectively. Although tll transcription is always rapidly induced, hkb converts graded ERK inputs into a variable time delay. Nuclei thus interpret ERK amplitude through the relative timing of tll and hkb transcription. Antagonistic regulatory paths acting on different timescales are hallmarks of an incoherent feedforward loop, which is sufficient to explain byn dynamics and adds temporal complexity to the steady-state model of byn stripe formation. We further show that 'blurring' of an all-or-none stimulus through intracellular diffusion non-locally produces a byn stripe. Overall, we provide a blueprint for using optogenetics to dissect developmental signal interpretation in space and time.

ERK and c-Myc signaling in host-derived tumor endothelial cells is essential for solid tumor growth.

The limited efficacy of the current antitumor microenvironment strategies is due in part to the poor understanding of the roles and relative contributions of the various tumor stromal cells to tumor development. Here, we describe a versatile in vivo anthrax toxin protein delivery system allowing for the unambiguous genetic evaluation of individual tumor stromal elements in cancer. Our reengineered tumor-selective anthrax toxin exhibits potent antiproliferative activity by disrupting ERK signaling in sensitive cells. Since this activity requires the surface expression of the capillary morphogenesis protein-2 (CMG2) toxin receptor, genetic manipulation of CMG2 expression using our cell-type-specific CMG2 transgenic mice allows us to specifically define the role of individual tumor stromal cell types in tumor development. Here, we established mice with CMG2 only expressed in tumor endothelial cells (ECs) and determined the specific contribution of tumor stromal ECs to the toxin's antitumor activity. Our results demonstrate that disruption of ERK signaling only within tumor ECs is sufficient to halt tumor growth. We discovered that c-Myc is a downstream effector of ERK signaling and that the MEK-ERK-c-Myc central metabolic axis in tumor ECs is essential for tumor progression. As such, disruption of ERK-c-Myc signaling in host-derived tumor ECs by our tumor-selective anthrax toxins explains their high efficacy in solid tumor therapy.

Intermittent ERK oscillations downstream of FGF in mouse embryonic stem cells.

Signal transduction networks generate characteristic dynamic activities to process extracellular signals and guide cell fate decisions such as to divide or differentiate. The differentiation of pluripotent cells is controlled by FGF/ERK signaling. However, only a few studies have addressed the dynamic activity of the FGF/ERK signaling network in pluripotent cells at high time resolution. Here, we use live cell sensors in wild-type and Fgf4-mutant mouse embryonic stem cells to measure dynamic ERK activity in single cells, for defined ligand concentrations and differentiation states. These sensors reveal pulses of ERK activity. Pulsing patterns are heterogeneous between individual cells. Consecutive pulse sequences occur more frequently than expected from simple stochastic models. Sequences become more prevalent with higher ligand concentration, but are rarer in more differentiated cells. Our results suggest that FGF/ERK signaling operates in the vicinity of a transition point between oscillatory and non-oscillatory dynamics in embryonic stem cells. The resulting heterogeneous dynamic signaling activities add a new dimension to cellular heterogeneity that may be linked to divergent fate decisions in stem cell cultures.

NLRP1 restricts porcine deltacoronavirus infection via IL-11 inhibiting the phosphorylation of the ERK signaling pathway.

Continuously emerging highly pathogenic coronaviruses remain a major threat to human and animal health. Porcine deltacoronavirus (PDCoV) is a newly emerging enterotropic swine coronavirus that causes large-scale outbreaks of severe diarrhea disease in piglets. Unlike other porcine coronaviruses, PDCoV has a wide range of species tissue tropism, including primary human cells, which poses a significant risk of cross-species transmission. Nucleotide-binding oligomerization domain-like receptor (NLR) family pyrin domain-containing 1 (NLRP1) has a key role in linking host innate immunity to microbes and the regulation of inflammatory pathways. We now report a role for NLRP1 in the control of PDCoV infection. Overexpression of NLRP1 remarkably suppressed PDCoV infection, whereas knockout of NLRP1 led to a significant increase in PDCoV replication. A mechanistic study revealed that NLRP1 suppressed PDCoV replication in cells by upregulating IL-11 expression, which in turn inhibited the phosphorylation of the ERK signaling pathway. Furthermore, the ERK phosphorylation inhibitor U0126 effectively hindered PDCoV replication in pigs. Together, our results demonstrated that NLRP1 exerted an anti-PDCoV effect by IL-11-mediated inhibition of the phosphorylation of the ERK signaling pathway, providing a novel antiviral signal axis of NLRP1-IL-11-ERK. This study expands our understanding of the regulatory network of NLRP1 in the host defense against virus infection and provides a new insight into the treatment of coronaviruses and the development of corresponding drugs.IMPORTANCECoronavirus, which mainly infects gastrointestinal and respiratory epithelial cells in vivo, poses a huge threat to both humans and animals. Although porcine deltacoronavirus (PDCoV) is known to primarily cause fatal diarrhea in piglets, reports detected in plasma samples from Haitian children emphasize the potential risk of animal-to-human spillover. Finding effective therapeutics against coronaviruses is crucial for controlling viral infection. Nucleotide-binding oligomerization-like receptor (NLR) family pyrin domain-containing 1 (NLRP1), a key regulatory factor in the innate immune system, is highly expressed in epithelial cells and associated with the pathogenesis of viruses. We demonstrate here that NLRP1 inhibits the infection of the intestinal coronavirus PDCoV through IL-11-mediated phosphorylation inhibition of the ERK signaling pathway. Furthermore, the ERK phosphorylation inhibitor can control the infection of PDCoV in pigs. Our study emphasizes the importance of NLRP1 as an immune regulatory factor and may open up new avenues for the treatment of coronavirus infection.

CircPCNXL2 promotes tumor growth and metastasis by interacting with STRAP to regulate ERK signaling in intrahepatic cholangiocarcinoma.

BACKGROUND: circular RNAs (circRNAs) have been reported to exert important effects in the progression of numerous cancers. However, the functions of circRNAs in intrahepatic cholangiocarcinoma (ICC) are still unclear. METHODS: circPCNXL2 (has_circ_0016956) were identified in paired ICC by circRNA microarray. Then, we assessed the biological functions of circPCNXL2 by CCK8, EdU, clone formation, transwell, wound healing assays, and xenograft models. RNA pull-down, mass spectrometry, and RNA immunoprecipitation (RIP) were applied to explore the interaction between cirrcPCNXL2 and serine-threonine kinase receptor-associated protein (STRAP). RNA pull-down, RIP and luciferase reporter assays were used to investigate the sponge functions of circPCNXL2. In the end, we explore the effects of circPCNXL2 and trametinib (a MEK1/2 inhibitor) in vivo. RESULTS: circPCNXL2 was upregulated in ICC tissues and cell lines, which promoted the proliferation and metastasis of ICC in vitro and in vivo. In terms of the mechanisms, circPCNXL2 could directly bind to STRAP and induce the interaction between STRAP and MEK1/2, resulting in the tumor promotion in ICC by activation of ERK/MAPK pathways. Besides, circPCNXL2 could regulate the expression of SRSF1 by sponging miR-766-3p and subsequently facilitated the growth of ICC. Finally, circPCNXL2 could partially inhibit the anti-tumor activity of trametinib in vivo. CONCLUSION: circPCNXL2 played a crucial role in the progression of ICC by interacting with STRAP to activate the ERK signaling pathway, as well as by modulating the miR-766-3p/SRSF1 axis. These findings suggest that circPCNXL2 may be a promising biomarker and therapeutic target for ICC.

PLA plastic particles disrupt bile acid metabolism leading to hepatic inflammatory injury in male mice.

Microplastics, such as polylactic acid (PLA), are ubiquitous environmental pollutants with unclear implications for health impact. This study aims to elucidate the mechanisms of PLA-induced inflammatory liver injury, focusing on disturbance of bile acid metabolism. The in vitro PLA exposure experiment was conducted using HepG2 cells to assess cell viability, cytokine secretion, and effects on bile acid metabolism. In vivo, male C57BL/6 J mice were exposed to PLA for ten days continuously, liver function and histopathological assessment were evaluated after the mice sacrificed. Molecular analyses including quantitative reverse transcription polymerase chain reaction (RT-qPCR) and Western blotting, were applied to evaluate the expression of bile acid metabolizing enzymes and transporters. PLA exposure resulted in decreased cell viability in HepG2 cells, increased inflammation and altered bile acid metabolism. In mice, PLA exposure resulted in decreased body weight and food intake, impaired liver function, increased hepatic inflammation, altered bile acid profiles, and dysregulated expression of bile acid metabolic pathways. PLA exposure disrupts bile acid metabolism through inhibition of the CYP7A1 enzyme and activation of the FGF-JNK/ERK signaling pathway, contributing to liver injury. These findings highlight the potential hepatotoxic effects of environmentally friendly plastics PLA and underscore the need for further research on their biological impact.

Cancer-associated fibroblasts promote stemness maintenance and gemcitabine resistance via HIF-1α/miR-21 axis under hypoxic conditions in pancreatic cancer.

Gemcitabine (GEM) resistance affects chemotherapy efficacy of pancreatic cancer (PC). Cancer-associated fibroblasts (CAFs) possess the ability of regulating chemoresistance. This study probed the mechanism of hypoxia-treated CAFs regulating cell stemness and GEM resistance in PC. Miapaca-2/SW1990 were co-cultured with PC-derived CAFs under normoxic/hypoxic conditions. Cell viability/self-renewal ability was determined by MTT/sphere formation assays, respectively. Protein levels of CD44, CD133, Oct4, and Sox2 were determined by western blot. GEM tumoricidal assay was performed. PC cell GEM resistance was evaluated by MTT assay. CAFs were cultured at normoxia/hypoxia. HIF-1α and miR-21 expression levels were assessed by RT-qPCR and western blot, with their binding sites and binding relationship predicted and verified. CAF-extracellular vesicles (EVs) were incubated with Miapaca-2 cells. The RAS/AKT/ERK pathway activation was detected by western blot. PC xenograft models were established and treated with hypoxic CAF-EVs and GEM. CAFs and PC cell co-culture increased cell stemness maintenance, GEM resistance, cell viability, stem cell sphere number, and protein levels of CD44, CD133, Oct4, and Sox2, and weakened GEM tumoricidal ability to PC cells, with the effects further enhanced by hypoxia. Hypoxia induced HIF-1α and miR-21 overexpression in CAFs. Hypoxia promoted CAFs to secrete high-level miR-21 EVs via the HIF-1α/miR-21 axis, and activated the miR-21/RAS/AKT/ERK pathway. CAF-EVs promoted GEM resistance in PC via the miR-21/RAS/ATK/ERK pathway in vivo. Hypoxia promoted CAFs to secrete high-level miR-21 EVs through the HIF-1α/miR-21 axis, and activated the miR-21/RAS/AKT/ERK pathway via EVs to trigger stemness maintenance and GEM resistance in PC.

Interference with MTHFD2 induces ferroptosis in ovarian cancer cells through ERK signaling to suppress tumor malignant progression.

Ovarian cancer (OC) is a deadliest gynecological cancer with the highest mortality rate. Methylenetetrahydrofolate dehydrogenase 2 (MTHFD2), a crucial tumor-promoting factor, is over-expressed in several malignancies including OC. The present study aimed to explore the role and mechanisms of MTHFD2 in OC malignant progression. Thus, cell proliferation, cycling, apoptosis, migration, and invasion were evaluated by CCK-8 assay, EdU assay, flow cytometry, wound healing, transwell assay and western blotting. Additionally, glycolysis was assessed by measuring the level of glucose and lactate production, as well as the expressions of GLUT1, HK2 and PKM2. Then the expression of ferroptosis-related proteins and ERK signaling was detected using western blotting. Ferroptosis was detected through the measurement of iron level, GSH, MDA and ROS activities. The results revealed that MTHFD2 was highly expressed in OC cells. Besides, interference with MTHFD2 induced ferroptosis, promoted ROS accumulation, destroyed mitochondrial function, reduced ATP content and inhibited glycolysis in OC cells. Subsequently, we further found that interference with MTHFD2 affected mitochondrial function and glycolysis in OC cells through ERK signaling. Moreover, interference with MTHFD2 affected ferroptosis to inhibit the malignant progression of OC cells. Collectively, our present study disclosed that interference with MTHFD2 induced ferroptosis in OC to inhibit tumor malignant progression through regulating ERK signaling.

Peroxiredoxin 3 regulates breast cancer progression via ERK-mediated MMP-1 expression.

Peroxiredoxin 3 (PRDX3), a mitochondrial hydrogen peroxide scavenger, is known to be upregulated during tumorigenesis and cancer progression. In this study, we provide evidence for the first time that PRDX3 could regulate cellular signaling pathways associated with Matrix Metalloproteinase-1 (MMP-1) expression and activity in breast cancer progression. We show that shRNA-mediated gene silencing of PRDX3 inhibits cell migration and invasion in two triple-negative breast cancer cell lines. Reciprocal experiments show that PRDX3 overexpression promotes invasion and migration of the cancer cells, processes which are important in the metastatic cascade. Notably, this phenomenon may be attributed to the activation of MMP-1, which is observed to be upregulated by PRDX3 in the breast cancer cells. Moreover, immunohistochemical staining of breast cancer tissues revealed a positive correlation between PRDX3 and MMP-1 expression in both epithelial and stromal parts of the tissues. Further pathway reporter array and luciferase assay demonstrated that activation of ERK signaling is responsible for the transcriptional activation of MMP-1 in PRDX3-overexpressed cells. These findings suggest that PRDX3 could mediate cancer spread via ERK-mediated activation of MMP-1. Targeted inhibition of ERK signaling may be able to inhibit tumor metastasis in triple-negative breast cancer.

Tenascin C as a novel zinc finger protein 750 target regulating the immunogenicity via DNA damage in lung squamous cell carcinoma.

Modulation of DNA damage repair in lung squamous cell carcinoma (LUSC) can result in the generation of neoantigens and heightened immunogenicity. Therefore, understanding DNA damage repair mechanisms holds significant clinical relevance for identifying targets for immunotherapy and devising therapeutic strategies. Our research has unveiled that the tumor suppressor zinc finger protein 750 (ZNF750) in LUSC binds to the promoter region of tenascin C (TNC), leading to reduced TNC expression. This modulation may impact the malignant behavior of tumor cells and is associated with patient prognosis. Additionally, single-cell RNA sequencing (scRNA-seq) of LUSC tissues has demonstrated an inverse correlation between ZNF750/TNC expression levels and immunogenicity. Manipulation of the ZNF750-TNC axis in vitro within LUSC cells has shown differential sensitivity to CD8+ cells, underscoring its pivotal role in regulating cellular immunogenicity. Further transcriptome sequencing analysis, DNA damage repair assay, and single-strand break analyses have revealed the involvement of the ZNF750-TNC axis in determining the preference for homologous recombination (HR) repair or non-homologous end joining (NHEJ) repair of DNA damage. with involvement of the Hippo/ERK signaling pathway. In summary, this study sheds light on the ZNF750-TNC axis's role in DNA damage repair regulation in LUSC, laying a groundwork for future translational research in immune cell therapy for LUSC.

DUSP1/MKP-1 represents another piece in the P2X7R intracellular signaling puzzle in cerebellar cells: our last journey with Mª Teresa along the purinergic pathways of Eden.

The P2X7 receptor (P2X7R) stands out within the purinergic family as it has exclusive pharmacological and regulatory features, and it fulfills distinct roles depending on the type of stimulation and cellular environment. Tonic activation of P2X7R promotes cell proliferation, whereas sustained activation is associated with cell death. Yet strikingly, prolonged P2X7R activation in rat cerebellar granule neurons and astrocytes does not affect cell survival. The intracellular pathways activated by P2X7Rs involve proteins like MAPKs, ERK1/2 and p38, and interactions with growth factor receptors could explain their behavior in populations of rat cerebellar cells. In this study, we set out to characterize the intracellular mechanisms through which P2X7Rs and Trk receptors, EGFR (epidermal growth factor receptor) and BDNFR (brain-derived neurotrophic factor receptor), regulate the dual-specificity phosphatase DUSP1. In cerebellar astrocytes, the regulation of DUSP1 expression by P2X7R depends on ERK and p38 activation. EGFR stimulation can also induce DUSP1 expression, albeit less strongly than P2X7R. Conversely, EGF was virtually ineffective in regulating DUSP1 in granule neurons, a cell type in which BDNF is the main regulator of DUSP1 expression and P2X7R only induces a mild response. Indeed, the regulation of DUSP1 elicited by BDNF reflects the balance between both transcriptional and post-transcriptional mechanisms. Importantly, when the regulation of DUSP1 expression is compromised, the viability of both astrocytes and neurons is impaired, suggesting this phosphatase is essential to maintain proper cell cytoarchitecture and functioning.

Quercetin conjugated PSC-derived exosomes to inhibit intimal hyperplasia via modulating the ERK, Akt, and NF-κB signaling pathways in the rat carotid artery post balloon injury.

The primary challenge in percutaneous coronary interventions for vascular restenosis is the occurrence of restenosis, which is defined by the excessive proliferation of neointimal tissue. Herein, our research team suggests that exosomes obtained from PSC, when paired with quercetin (Q@PSC-E), successfully reduce neointimal hyperplasia in a Sprague-Dawley rat model. Furthermore, the physical properties of the synthesized Q@PSC-E were examined using UV-vis, DLS, and FT-IR characterization techniques. The rats were subjected to balloon injury (BI) utilizing a 2-Fr Fogarty arterial embolectomy balloon catheter. Intimal hyperplasia and the degree of VSMC proliferation were evaluated using histological analysis in the rat groups that received a dosage of Q@PSC-E at 30 mg/kg/d. Significantly, Q@PSC-E inhibited cell proliferation through a pathway that does not include lipoxygenase, as demonstrated by [3H] thymidine incorporation, MTT, and flow cytometry studies. Additionally, the data indicate that Q@PSC-E hinders cell proliferation by targeting particular events that promote cell growth, including the activation of Akt and NF-κB, disruption of cell-cycle progression and also obstructs the ERK signaling pathway.

GABRP Promotes the Metastasis of Pancreatic Cancer by Activation of the MEK/ERK Signaling Pathway.

Pancreatic cancer remains the common cancer with the worst prognosis because of its late diagnosis and extensive metastasis. This study aimed to investigate the effects of GABRP on pancreatic cancer metastasis and the molecular mechanism. The expression of GABRP was measured using the quantitative real-time PCR and western blot. The biological behaviors of cancer cells were assessed using the cell counting kit-8, Transwell assay, and western blot. The regulation of GABRP on the MEK/ERK pathway was detected by western blot. The results indicated that GABRP was overexpressed in pancreatic cancer tissues and cells. Knockdown of GABRP suppressed cell viability, invasion, migration, and epithelial-mesenchymal transition (EMT), whereas GABRP overexpression facilitated these biological behaviors. Inactivation of the MEK/ERK pathway reversed the effects on cellular processes induced by GABRP. Moreover, silencing of GABRP inhibited tumor growth. In conclusion, GABRP promoted the progression of pancreatic cancer by facilitating cell metastasis and tumor growth via activating the MEK/ERK pathway. The findings suggest that GABRP has the potential to be a therapeutic target for the metastatic pancreatic cancer.

Resveratrol protects mesangial cells under high glucose by regulating the miR-1231/IGF1/ERK pathway.

Diabetic nephropathy (DN) is one of the complications of diabetes mellitus and the main cause of end-stage renal disease (ESRD), which is a serious threat to human health. In DN, mesangial cells (MCs) are a critical target cell that perform a variety of key functions, and abnormal proliferation of MCs is a common and prominent pathological change in DN. In recent years, the investigation of Chinese medicine interventions for DN has increased significantly in recent years due to the many potential adverse effects and controversies associated with the treatment of DN with Western medicines. In this study, we evaluated the protective effect of resveratrol (RES), an active ingredient known as a natural antioxidant, on HMCs under high glucose and explored its possible mechanism of action. We found that RES inhibited the proliferation of human mesangial cell (HMC) under high glucose and blocked cell cycle progression. In the high glucose environment, RES upregulated miR-1231, reduced IGF1 expression, inhibited the activity of the extracellular signal-regulated kinase (ERK) signaling pathway and reduced levels of the inflammatory factors TNF-α and IL-6. In addition, we found that miR-1231 mimics were synergistically inhibited with RES, whereas miR-1231 inhibitor attenuated the protective effect of RES on HMCs. Thus, our results suggest that the protective effect of RES on HMCs under high glucose is achieved, at least in part, through modulation of the miR-1231/IGF1/ERK pathway. The discovery of this potential mechanism may provide a new molecular therapeutic target for the prevention and treatment of DN, and may also bring new ideas for the clinical research in DN.

Identification of cell type-specific correlations between ERK activity and cell viability upon treatment with ERK1/2 inhibitors.

Increased MAPK signaling is a hallmark of various cancers and is a central regulator of cell survival. Direct ERK1/2 inhibition is considered a promising approach to avoid ERK1/2 reactivation caused by upstream kinases BRAF, MEK1/2, and KRAS, as well as by receptor tyrosine kinase inhibitors, but the dynamics and selectivity of ERK1/2 inhibitors are much less studied compared with BRAF or MEK inhibitors. Using ERK1/2 and downstream kinase ELK1 reporter cell lines of lung cancer (H1299; NRASQ61K), colon cancer (HCT-116; KRASG13D), neuroblastoma (SH-SY5Y), and leukemia (U937), we examined the relationship between ERK inhibition and drug-induced toxicity for five ERK inhibitors: SCH772984, ravoxertinib, LY3214996, ulixertinib, and VX-11e, as well as one MEK inhibitor, PD0325901. Comparing cell viability and ERK inhibition revealed different ERK dependencies for these cell lines. We identify several drugs, such as SCH772984 and VX-11e, which induce excessive toxicity not directly related to ERK1/2 inhibition in specific cell lines. We also show that PD0325901, LY3214996, and ulixertinib are prone to ERK1/2 reactivation over time. We distinguished two types of ERK1/2 reactivation: the first could be reversed by adding a fresh dose of inhibitors, while the second persists even after additional treatments. We also showed that cells that became resistant to the MEK1/2 inhibitor PD0325901 due to ERK1/2 reactivation remained sensitive to ERK1/2 inhibitor ulixertinib. Our data indicate that correlation of ERK inhibition with drug-induced toxicity in multiple cell lines may help to find more selective and effective ERK1/2 inhibitors.

ERK mediates interferon gamma-induced melanoma cell death.

BACKGROUND: Interferon-gamma (IFNγ) exerts potent growth inhibitory effects on a wide range of cancer cells through unknown signaling pathways. We pursued complementary screening approaches to characterize the growth inhibition pathway. METHODS: We performed chemical genomics and whole genome targeting CRISPR/Cas9 screens using patient-derived melanoma lines to uncover essential nodes in the IFNγ-mediated growth inhibition pathway. We used transcriptomic profiling to identify cell death pathways activated upon IFNγ exposure. Live imaging experiments coupled with apoptosis assays confirmed the involvement of these pathways in IFNγ-mediated cell death. RESULTS: We show that IFNγ signaling activated ERK. Blocking ERK activation rescued IFNγ-mediated apoptosis in 17 of 23 (~ 74%) cell lines representing BRAF, NRAS, NF1 mutant, and triple wild type subtypes of cutaneous melanoma. ERK signaling induced a stress response, ultimately leading to apoptosis through the activity of DR5 and NOXA proteins. CONCLUSIONS: Our results provide a new understanding of the IFNγ growth inhibition pathway, which will be crucial in defining mechanisms of immunotherapy response and resistance.

BRAF(V600E) mutation together with loss of Trp53 or pTEN drives the origination of hairy cell leukemia from B-lymphocytes.

Hairy cell leukemia (HCL) is a B-lymphoma induced by BRAF(V600E) mutation. However, introducing BRAF(V600E) in B-lymphocytes fails to induce hematological malignancy, suggesting that BRAF(V600E) needs concurrent mutations to drive HCL ontogeny. To resolve this issue, here we surveyed human HCL genomic sequencing data. Together with previous reports, we speculated that the tumor suppressor TP53, P27, or PTEN restrict the oncogenicity of BRAF(V600E) in B-lymphocytes, and therefore that their loss-of-function facilitates BRAF(V600E)-driven HCL ontogeny. Using genetically modified mouse models, we demonstrate that indeed BRAF(V600E)KI together with Trp53KO or pTENKO in B-lymphocytes induces chronic lymphoma with pathological features of human HCL. To further understand the cellular programs essential for HCL ontogeny, we profiled the gene expression of leukemic cells isolated from BRAF(V600E)KI and Trp53KO or pTENKO mice, and found that they had similar but different gene expression signatures that resemble that of M2 or M1 macrophages. In addition, we examined the expression signature of transcription factors/regulators required for germinal center reaction and memory B cell versus plasma cell differentiation in these leukemic cells and found that most transcription factors/regulators essential for these programs were severely inhibited, illustrating why hairy cells are arrested at a transitional stage between activated B cells and memory B cells. Together, our study has uncovered concurrent mutations required for HCL ontogeny, revealed the B cell origin of hairy cells and investigated the molecular basis underlying the unique pathological features of the disease, with important implications for HCL research and treatment.

Aberrant cytoplasmic localization of ARID1B activates ERK signaling and promotes oncogenesis.

The ARID1B (BAF250b) subunit of the human SWI/SNF chromatin remodeling complex is a canonical nuclear tumor suppressor. We employed in silico prediction, intracellular fluorescence and cellular fractionation-based subcellular localization analyses to identify the ARID1B nuclear localization signal (NLS). A cytoplasm-restricted ARID1B-NLS mutant was significantly compromised in its canonical transcription activation and tumor suppressive functions, as expected. Surprisingly however, cytoplasmic localization appeared to induce a gain of oncogenic function for ARID1B, as evidenced from several cell line- and mouse xenograft-based assays. Mechanistically, cytoplasm-localized ARID1B could bind c-RAF (RAF1) and PPP1CA causing stimulation of RAF-ERK signaling and ß-catenin (CTNNB1) transcription activity. ARID1B harboring NLS mutations derived from tumor samples also exhibited aberrant cytoplasmic localization and acquired a neo-morphic oncogenic function via activation of RAF-ERK signaling. Furthermore, immunohistochemistry on a tissue microarray revealed significant correlation of ARID1B cytoplasmic localization with increased levels of active forms of ERK1 and ERK2 (also known as MAPK3 and MAPK1) and of ß-catenin, as well as with advanced tumor stage and lymph node positivity in human primary pancreatic tumor tissues. ARID1B therefore promotes oncogenesis through cytoplasm-based gain-of-function mechanisms in addition to dysregulation in the nucleus.This article has an associated First Person interview with the first author of the paper.

CMTM3 deficiency induces cardiac hypertrophy by regulating MAPK/ERK signaling.

OBJECTIVES: Cardiac hypertrophy is the heart's compensatory response stimulated by various pathophysiological factors. However, prolonged cardiac hypertrophy poses a significant risk of progression to heart failure, lethal arrhythmias, and even sudden cardiac death. For this reason, it is crucial to effectively prevent the occurrence and development of cardiac hypertrophy. CMTM is a superfamily of human chemotaxis, which is involved in immune response and tumorigenesis. CMTM3 expressed widely in tissues, including the heart, but its cardiac function remains unclear. This research aims to explore the effect and mechanism of CMTM3 in the development of cardiac hypertrophy. METHODS AND RESULTS: We generated a Cmtm3 knockout mouse model (Cmtm3-/-) as the loss-of-function approach. CMTM3 deficiency induced cardiac hypertrophy and further exacerbated hypertrophy and cardiac dysfunction stimulated by Angiotensin Ⅱ infusion. In Ang Ⅱ-infusion stimulated hypertrophic hearts and phenylephrine-induced hypertrophic neonatal cardiomyocytes, CMTM3 expression significantly increased. However, adenovirus-mediated overexpression of CMTM3 inhibited the hypertrophy of rat neonatal cardiomyocytes induced by PE stimulation. In terms of mechanism, RNA-seq data revealed that Cmtm3 knockout-induced cardiac hypertrophy was related to MAPK/ERK activation. In vitro, CMTM3 overexpression significantly inhibited the increased phosphorylation of p38 and ERK induced by PE stimulation. CONCLUSIONS: CMTM3 deficiency induces cardiac hypertrophy and aggravates hypertrophy and impaired cardiac function stimulated by angiotensin Ⅱ infusion. The expression of CMTM3 increases during cardiac hypertrophy, and the increased CMTM3 can inhibit further hypertrophy of cardiomyocytes by inhibiting MAPK signaling. Thus, CMTM3 plays a negative regulatory effect in the occurrence and development of cardiac hypertrophy.