Grb2-associated Binding Protein Research Articles

Association of GAB1 gene with asthma susceptibility and the efficacy of inhaled corticosteroids in children

Asthma is a polygenic disease that may onset during childhood. Inhaled corticosteroids (ICS) are the main therapy in asthma, although their efficacy varies among individuals. Nuclear factor κB (NF-κB) is an important target of ICS treatment of asthma. Recent research has reported that GRB2 associated binding protein 1 (GAB1) gene may participate in the pathogenesis of asthma by regulating the NF-κB pathway. Therefore, we used the technique of an improved multiplex ligation detection reaction to sequence GAB1 gene and investigated the involvement of Single-nucleotide variants (SNVs) in GAB1 gene in asthma and ICS efficacy in asthmatic children. We found no differences between asthma cases and controls in allele or genotype frequencies of GAB1. Haplotype analysis showed an increased tendency for AGGAGC frequency in asthma patients compared with controls (OR = 2.69, p = 0.018). The percentage of EOS and genotype distribution of rs1397527 were associated (p = 0.007). The EOS percentage was higher in GT genotype when compared to the GG genotype (5.50 vs 3.00, Bonferroni adjusted p = 0.005). After 12-weeks ICS treatment, GAB1 rs1397527 TT and GT genotype carriers had a smaller change in forced expiratory volume in 1 second/forced vital capacity (FEV1/FVC) than GG carriers (p = 0.009), and rs3805236 GG and AG genotype carriers also had a smaller change in FEV1/FVC than AA carriers (p = 0.025). For ICS response, the frequency of GG genotype of rs1397527 was significantly higher in good responders (p = 0.038). The generalized multifactor dimensionality reduction (GMDR) analysis showed a best significant four-order model (rs1397527, allergen exposure, environmental tobacco smoke exposure, and pet exposure) involving gene-environment interactions (p = 0.001). In summary, we found that GAB1 SNVs were not associated with asthma susceptibility. Haplotype AGGAGC was a risk factor for asthma. GAB1 variants were associated with eosinophils and ICS response in asthmatics. Furthermore, gene-environment interaction was observed.

Leaves and pseudostems extract of Curcuma longa attenuates immunoglobulin E/bovine serum albumin-stimulated bone marrow-derived cultured mast cell activation and passive cutaneous anaphylaxis in BALB/c mice

Ethnopharmacological relevanceCurcuma longa, known as turmeric, is an herbaceous perennial plant belonging to the genus Curcuma. It is dispersed throughout tropical and subtropical regions worldwide. Since ancient times, turmeric has been used as an ethnomedicinal plant in the Ayurvedic system, particularly in Asian countries. Rhizomes of turmeric possess several pharmacological properties that give high value as a medicinal remedy for treating a range of conditions, including inflammation, pain, allergies, and digestive issues. Moreover, turmeric leaves and pseudostems also contain a variety of health-enhancing secondary metabolites, such as curcumin, flavonoids, and other phenolic compounds, which exhibit anti-inflammatory, antitumor, antibacterial, and antioxidant properties. Aim of the studyAllergic diseases are a group of immune-mediated disorders mainly caused by an immunoglobulin E (IgE)-dependent immunological response to an innocuous allergen. Therefore, this study aimed to investigate the effect of leaves and pseudostems extract of turmeric (TLSWE-8510) on IgE/bovine serum albumin (BSA)-stimulated allergic responses in mouse bone marrow-derived cultured mast cells (BMCMCs) and passive cutaneous anaphylaxis (PCA) in BALB/c mice. Materials and methodsThe effect of TLSWE-8510 on mast cell degranulation has been evaluated by investigating the release of β-hexosaminidase and histamine in IgE/BSA-stimulated BMCMCs. Additionally, anti-allergic properties of TLSWE-8510 on IgE/BSA-stimulated BMCMCs were investigated using suppression of nuclear factor-kappa B (NF-κB), and spleen tyrosine kinase (Syk)-linker for T-cell activation (LAT)-extracellular-signal-regulated kinase (ERK)-GRB2 associated binding protein 2 (Gab2) signaling pathway and downregulation of allergy-related cytokines and chemokines expression. Furthermore, in vivo, studies were conducted using IgE-mediated PCA in BALB/c mice. ResultsTLSWE-8510 treatment significantly inhibited the degranulation of IgE/BSA-stimulated BMCMCs by inhibiting the release of β-hexosaminidase and histamine dose-dependently. Additionally, TLSWE-8510 reduced the expression of high-affinity IgE receptors (Fc epsilon receptor I-FcεRI) on the surface of BMCMCs and the binding of IgE to FcεRI. Besides, the expression of cytokines and chemokines is triggered by IgE/BSA stimulation via activating the allergy-related signaling pathways. TLSWE-8510 dose-dependently downregulated the mRNA expression and the production of allergy-related cytokines (interleukin (IL)-1β, IL-3, IL-4, IL-5, IL-6, IL-13, tumor necrosis factor (TNF)-α, and interferon (IFN)-γ), and chemokines (thymus and activation-regulated chemokine (TARC), and regulated upon activation, normal T cell expressed and secreted (RANTES)) by regulating the phosphorylation of downstream signaling molecules, NF-κB, and Syk, LAT, ERK and Gab2 in IgE/BSA-stimulated BMCMCs. Moreover, PCA reaction in IgE/BSA-stimulated BALB/c mice ears was effectively decreased by TLSWE-8510 treatment in a dose-dependent manner. ConclusionsThese results collectively demonstrated that TLSWE-8510 suppressed mast cell degranulation by inhibiting the release of chemical mediators related to allergies. TLSWE-8510 downregulated the allergy-related cytokines and chemokines expression and phosphorylation of downstream signaling molecules in IgE/BSA-stimulated BMCMCs. Furthermore, in vivo studies with IgE-mediated PCA reaction in the BALB/c mice ears were attenuated by TLSWE-8510 treatment. These findings revealed that TLSWE-8510 has the potential as a therapeutic agent for the treatment of allergic diseases.

Gab3 Plays an Essential Role in CD8 + T-Cell Expansion through Regulating IL-2-Mediated Activation of PI3K/AKT/mTOR and Erk/FoxO Pathways

Grb2-associated binding protein 3 (Gab3) is a member of the Gab family of scaffolding proteins. Its role in T cells is largely unknown. In the current study, we investigated its in vivo function in CD8 + T cells using the lymphocytic choriomeningitis virus (LCMV) Armstrong infection model. We created BM chimera mice with a specific deficiency of Gab3 in CD8 + T cells. This was achieved by transplanting a mixture of BM cells obtained from CD8-deficient mice and Gab3-deficient (Gab3 -/-) mice (1) into lethally irradiated CD45.1 mice. The control chimera mice were lethally irradiated CD45.1 mice transplanted with a mixture of BM cells from CD8-deficient mice and WT mice. Twelve weeks after the BM transplantation, the Gab3 -/- and control BM chimera mice were infected with Armstrong. On day eight following infection, Gab3 -/- CD8 + T cells exhibited reduced expansion compared to WT CD8 + T cells. Importantly, Gab3 -/- LCMV antigen specific CD8 + T cells, GP33 + and NP396 + CD8 + T cells, were significantly reduced compared to their WT counterparts. Furthermore, CD8 + T cells in Gab3 -/- BM chimera mice showed significant increase of apoptosis compared to that in WT BM chimera mice following LCMV infection. To explore the mechanism underlying the observed phenotype, we used a primary CD8 + T-cell culture system, in which we stimulated CD8 + T cells with anti-CD3 and anti-CD28 followed by expanding the cells in IL-2. We observed that Gab3 deficiency severely impaired IL-2-induced CD8 + T cell expansion. Gab3 -/- CD8 + T cells in this culture system displayed increased apoptosis and reduced proliferation compared to wild-type (WT) controls, which may account for the reduced CD8 + T cell expansion. We then performed Seahorse analysis of the cultured CD8 + T cells after IL-2 stimulation. We found that IL-2-mediated extracellular acidification rate (ECAR) and oxygen consumption rate (OCR) were largely impaired in Gab3 -/- CD8 + T cells compared to WT CD8 + T cells. This data indicates a crucial role of Gab3 in both glycolysis and mitochondrial respiration. We further investigated how Gab3 regulates the IL-2 signaling pathway by conducting transcriptomic and phosphoproteomic analysis. RNA-seq analysis revealed that genes associated with mTOR signaling, glucose metabolism, cell cycle, and Erk/MAPK targets were highly enriched in WT CD8 + T cells, whereas genes related to apoptosis, autophagy, and FoxO3 targets were highly enriched in Gab3 -/- CD8 + T cells. In contrast, the gene set enrichment analysis (GSEA) showed that IL-2-induced Stat5 target gene expression was comparable between WT and Gab3 -/- CD8 + T cells, indicating an important role of Gab3 specifically in the activation of the mTOR and Erk/MAPK pathways downstream of IL-2 signaling. Consistent with that observed in the transcriptomic analysis, phosphoproteomics analysis revealed a significant reduction in IL-2-induced phosphorylation of key components in ERK/FOXO signaling pathway, including RafA, MAPK1, FoxO3, AMPK, and mTOR downstream molecule Eif4B in Gab3 -/- CD8 + T cells compared to WT CD8 + cells. Furthermore, phosphorylation of Stat5 at different sites was comparable between Gab3 -/- and WT CD8 + T cells, further supporting an important role of Gab3 specifically in IL-2 mediated mTOR and Erk/MAPK activation. Finally, we validated the results obtained from the transcriptomic and phosphoproteomic analyses using western blot analysis. Our findings revealed that in Gab3 -/- CD8 + T cells, the IL-2-induced phosphorylation of signaling molecules in the mTOR pathway (S6K, S6, 4EBP1, AKT) and the Erk pathway (Erk) was impaired. Notably, the phosphorylation of Stat5 remained intact despite the absence of Gab3. In summary, our studies have demonstrated that Gab3 plays an essential role in CD8 + T-cell expansion by promoting cell proliferation and prohibiting cell apoptosis through regulating IL-2-mediated activation of PI3K/AKT/mTOR and Erk-FoxO3 pathways. (1) We gratefully acknowledge that Gab3-deficient mice were originally generated by Dr. Larry Rohrschneider at Fred Hutchinson Cancer Research Center ( Mol Cell Biol 2003 Apr;23(7):2415-24. doi: 10.1128/MCB.23.7.2415-2424.2003.). The mice were subsequently backcrossed to C57BL/6.

Upregulation of miR-200a improves ureteral obstruction-induced renal fibrosis via GAB1/Wnt/β-catenin signaling

BackgroundRenal fibrosis is a basic pathological change of almost all chronic kidney disorders. Epithelial–mesenchymal transition (EMT) and excessive extracellular matrix (ECM) accumulation play a crucial role in the process of fibrosis. MethodsWestern blot and qRT-PCR were accomplished to analyze the expression levels of target proteins and genes, respectively. The fibrotic levels in the renal tissues of rats were confirmed utilizing Masson staining. Expression of ECM-related α-SMA in the renal tissues was determined by immunohistochemistry assay. The combination of GRB2 associated binding protein 1 (GAB1) and miR-200a was ensured by starBase database and luciferase reporter assay. ResultsOur data uncovered that miR-200a was downregulated, but GAB1 was upregulated in the renal tissues of the rat experienced unilateral ureteral obstruction (UUO). Overexpression of miR-200a improved tissues fibrosis, suppressed GAB1 expression and ECM deposition, and inactivated Wnt/β-catenin in UUO rats. Moreover, miR-200a expression was inhibited, while GAB1 expression was facilitated in the TGF-β1-induced HK-2 cells. In TGF-β1-induced HK-2 cells, miR-200a overexpression inhibited GAB1 expression, also declined ECM-related proteins and mesenchymal markers expression. Oppositely, miR-200a overexpression facilitated epithelial marker expression in the TGF-β1-induced HK-2 cells. Next, the data revealed that miR-200a inhibited GAB1 expression through binding to the mRNA 3′-UTR of GAB1. Increasing of GAB1 reversed the regulation of miR-200a to GAB1 expression, Wnt/β-catenin signaling activation, EMT and ECM accumulation. ConclusionOverall, miR-200a increasing improved renal fibrosis through attenuating EMT and ECM accumulation by limiting Wnt/β-catenin signaling via sponging GAB1, indicating miR-200a may be a promising objective for renal disease therapy.

Upregulation of miR-200a improves ureteral obstruction-induced renal fibrosis via GAB1/Wnt/β-catenin signaling

BackgroundRenal fibrosis is a basic pathological change of almost all chronic kidney disorders. Epithelial–mesenchymal transition (EMT) and excessive extracellular matrix (ECM) accumulation play a crucial role in the process of fibrosis. MethodsWestern blot and qRT-PCR were accomplished to analyze the expression levels of target proteins and genes, respectively. The fibrotic levels in the renal tissues of rats were confirmed utilizing Masson staining. Expression of ECM-related α-SMA in the renal tissues was determined by immunohistochemistry assay. The combination of GRB2 associated binding protein 1 (GAB1) and miR-200a was ensured by starBase database and luciferase reporter assay. ResultsOur data uncovered that miR-200a was downregulated, but GAB1 was upregulated in the renal tissues of the rat experienced unilateral ureteral obstruction (UUO). Overexpression of miR-200a improved tissues fibrosis, suppressed GAB1 expression and ECM deposition, and inactivated Wnt/β-catenin in UUO rats. Moreover, miR-200a expression was inhibited, while GAB1 expression was facilitated in the TGF-β1-induced HK-2 cells. In TGF-β1-induced HK-2 cells, miR-200a overexpression inhibited GAB1 expression, also declined ECM-related proteins and mesenchymal markers expression. Oppositely, miR-200a overexpression facilitated epithelial marker expression in the TGF-β1-induced HK-2 cells. Next, the data revealed that miR-200a inhibited GAB1 expression through binding to the mRNA 3′-UTR of GAB1. Increasing of GAB1 reversed the regulation of miR-200a to GAB1 expression, Wnt/β-catenin signaling activation, EMT and ECM accumulation. ConclusionOverall, miR-200a increasing improved renal fibrosis through attenuating EMT and ECM accumulation by limiting Wnt/β-catenin signaling via sponging GAB1, indicating miR-200a may be a promising objective for renal disease therapy.

Clinicomorphological and molecular analysis of medulloblastoma and association with survival: A single tertiary care center experience.

Medulloblastoma (MB) is a heterogeneous disease, displaying distinct genetic profiles, with specific molecular subgroups. Various clinical, pathological and molecular variables have been associated with disease outcome and therefore utilised in risk stratification of patients. To perform molecular classification of medulloblastoma using surrogate immunohistochemistry (IHC) and associate molecular subgroups, histopathological types, and available clinicopathological parameters with overall survival (OS) of MB patients. This study included 65 medulloblastoma patients. Immunohistochemical staining, using β-catenin YAP1 and GRB2-Associated Binding Protein 1 (GAB1) antibodies was used to classify MB cases into wingless signalling (WNT) activated, sonic hedgehog (SHH) activated, and non-WNT/non-SHH molecular subgroups. The relevant statistical analysis was done using GraphPad Prism version 9.3.0. Histological patterns included classic (40 cases, 62%), desmoplastic nodular (D/N) (14 cases, 22%), large cell/anaplastic (LC/A) (9 cases, 13%), medulloblastoma with extensive nodularity (MBEN) (1 case, 1.5%) and one special subtype, i.e., medulloblastoma with myogenic and melanotic differentiation. Molecular subgroups included WNT (4 cases, 6%), SHH (34 cases, 52%), and non-WNT/non-SHH (27 cases, 42%) subgroups. Histopathological types differed significantly according to tumor location, degree of anaplasia and molecular subgroups. Molecular subgroups differed significantly in age distribution and tumor location. The probability of survival was 78% and 68% after 1 and 2 years, respectively. Infants (<3 years of age), LC/A pattern, and TP53-mutant status among SHH subgroup conferred poor prognosis in our study. At the end of the study (at 65 months of maximum follow-up period) probability of survival was 51%. Immunohistochemical analysis helps in molecular classification of medulloblastoma in majority of the cases as well as helps in predicting prognosis and treatment response.

Gαi1/3 mediate Netrin-1-CD146-activated signaling and angiogenesis.

Netrin-1 binds to the high-affinity receptor CD146 to activate downstream signaling and angiogenesis. Here, we examine the role and underlying mechanisms of G protein subunit alpha i1 (Gαi1) and Gαi3 in Netrin-1-induced signaling and pro-angiogenic activity. In mouse embryonic fibroblasts (MEFs) and endothelial cells, Netrin-1-induced Akt-mTOR (mammalian target of rapamycin) and Erk activation was largely inhibited by silencing or knockout of Gαi1/3, whereas signaling was augmented following Gαi1/3 overexpression. Netrin-1 induced Gαi1/3 association with CD146, required for CD146 internalization, Gab1 (Grb2 associated binding protein 1) recruitment and downstream Akt-mTOR and Erk activation. Netrin-1-induced signaling was inhibited by CD146 silencing, Gab1 knockout, or Gαi1/3 dominant negative mutants. Netrin-1-induced human umbilical vein endothelial cell (HUVEC) proliferation, migration and tube formation were inhibited by Gαi1/3 short hairpin RNA (shRNA), but were potentiated by ectopic Gαi1/3 overexpression. In vivo, intravitreous injection of Netrin-1 shRNA adeno-associated virus (AAV) significantly inhibited Akt-mTOR and Erk activation in murine retinal tissues and reduced retinal angiogenesis. Endothelial knockdown of Gαi1/3 significantly inhibited Netrin1-induced signaling and retinal angiogenesis in mice. Netrin-1 mRNA and protein expression were significantly elevated in retinal tissues of diabetic retinopathy (DR) mice. Importantly, silence of Netrin-1, by intravitreous Netrin-1 shRNA AAV injection, inhibited Akt-Erk activation, pathological retinal angiogenesis and retinal ganglion cells degeneration in DR mice. Lastly, Netrin-1 and CD146 expression is significantly increased in the proliferative retinal tissues of human proliferative diabetic retinopathy patients. Together, Netrin-1 induces CD146-Gαi1/3-Gab1 complex formation to mediate downstream Akt-mTOR and Erk activation, important for angiogenesis in vitro and in vivo.

High expression of GRB2 associated binding protein 3 mRNA predicts positive prognosis in melanoma.

Malignant melanoma is the most aggressive form of skin cancer, and it is characterized by poor prognosis in patients with metastatic diseases. Accurate prediction of prognosis is crucial for therapeutic decisions. In this study, bioinformatics analysis was used to explore the prognostic value of growth factor receptor-bound protein 2-associated binding protein 3 (GAB3) mRNA. RNA transcriptome sequencing data and clinical data from The Cancer Genome Atlas and genotype-tissue expression (GTEx) were analyzed for differentially expressed genes in high and low GAB3 mRNA expression groups in melanoma. Performing gene enrichment analysis and constructing protein-protein interaction networks. High expression of GAB3 was significantly correlated with a lower T stage, melanoma Clark level, Breslow depth, and melanoma ulceration. And high GAB3 expression was also associated with better progression-free interval in T1 and T2 stages and N0 stage and longer overall survival in T1 and T2 stages, N0 stage, and N1 stage. GAB3 promoted high levels of infiltration of macrophages and activated natural killer cells in melanoma. High expression of GAB3 predicted a positive prognosis in early-stage melanoma that may be mediated by the anticancer immune response.

Gab1 Overexpression Attenuates Susceptibility to Ventricular Arrhythmias in Pressure Overloaded Heart Failure Mouse Hearts.

Grb2 associated binding protein 1 (Gab1) is an adaptor protein that is important for intracellular signal transduction which involved in several pathological process. However, the role of Gab1 in pressure overload-induced ventricular arrhythmias (VAs) remain poorly understood. In the current study, we aimed to test the role of Gab1 in VA susceptibility induced by pressure overload. We overexpressed Gab1 in the hearts using an adeno-associated virus 9 (AAV9) system through tail vein injection. Aortic banding (AB) surgery was performed in C57BL6/J mice to induce heart failure (HF). Four weeks following AB, histology, echocardiography, and biochemical analysis were conducted to investigate cardiac structural remodeling and electrophysiological studies were performed to check the electrical remodeling. Western blot analysis was used to explore the underlying mechanisms. The mRNA and protein expression were downregulated in AB hearts compared to sham hearts. Gab1 overexpression significantly reversed AB-induced cardiac structural remodeling including ameliorated AB-induced cardiac dysfunction, cardiac fibrosis, and inflammatory response. Moreover, Gab1 overexpression also markedly alleviated AB-induced electrical remodeling including ion channel alterations and VA susceptibility. Mechanistically, we found that TLR4/MyD88/NF-κB contributes to the cardio protective effect of Gab1 overexpression on AB-induced VAs. Our study manifested that Gab1 may serve as a promising anti-arrhythmic target via inhibiting TLR4/MyD88/NF-κB signaling pathway induced by AB.

Receptor-kinase EGFR-MAPK adaptor proteins mediate the epithelial response to Candida albicans via the cytolytic peptide toxin, candidalysin

Candida albicans (C. albicans) is a dimorphic commensal human fungal pathogen that can cause severe oropharyngeal candidiasis (oral thrush) in susceptible hosts. During invasive infection, C. albicans hyphae invade oral epithelial cells (OECs) and secrete candidalysin, a pore-forming cytolytic peptide that is required for C. albicans pathogenesis at mucosal surfaces. Candidalysin is produced in the hyphal invasion pocket and triggers cell damage responses in OECs. Candidalysin also activates multiple MAPK-based signaling events that collectively drive the production of downstream inflammatory mediators that coordinate downstream innate and adaptive immune responses. The activities of candidalysin are dependent on signaling through the epidermal growth factor receptor (EGFR). Here, we interrogated known EGFR-MAPK signaling intermediates for their roles mediating the OEC response to C. albicans infection. Using RNA silencing and pharmacological inhibition, we identified five key adaptors, including growth factor receptor-bound protein 2 (Grb2), Grb2-associated binding protein 1 (Gab1), Src homology and collagen (Shc), SH2-containing protein tyrosine phosphatase-2 (Shp2), and casitas B-lineage lymphoma (c-Cbl). We determined that all of these signaling effectors were inducibly phosphorylated in response to C. albicans. These phosphorylation events occurred in a candidalysin-dependent manner and additionally required EGFR phosphorylation, matrix metalloproteinases (MMPs), and cellular calcium flux to activate a complete OEC response to fungal infection. Of these, Gab1, Grb2, and Shp2 were the dominant drivers of ERK1/2 activation and the subsequent production of downstream innate-acting cytokines. Together, these results identify the key adaptor proteins that drive the EGFR signaling mechanisms that underlie oral epithelial responses to C. albicans.

Inhibition of GAB2 expression has a protective effect on osteoarthritis:An in vitro and in vivo study

Osteoarthritis is a chronic age-related degenerative disease associated with varying degrees of pain and joint mobility disorders. Grb2-associated-Binding protein-2 (GAB2) is an intermediate molecule that plays a role downstream in a variety of signaling pathways, such as inflammatory signaling pathways. The role of GAB2 in the pathogenesis of OA has not been fully studied. In this study, we found that GAB2 expression was elevated in chondrocytes after constructing in vivo and in vitro models of OA. Inhibition of GAB2 by siRNA decreased the expression of MMP3, MMP13, iNOS, COX2, p62, and increased the expression of COL2, SOX9, ATG7, Beclin-1 and LC3II/LC3I. Furthermore, inhibition of GAB2 expression inhibited interleukin-1β (IL-1β) -induced mitogen-activated protein kinase (MAPK) and nuclear factor κB (NF-κB) signaling. In vivo studies, we found that reduced GAB2 expression effectively delayed cartilage destruction in a mouse model of OA induced by destabilisation of the medial meniscus (DMM). In conclusion, our study demonstrates that GAB2 is a potential therapeutic target for OA.

Association of GAB2 with Quality of Life and Negative Emotions in Patients with Gastric Cancer after Postoperative Comprehensive Care.

GRB2-associated binding protein 2 (GAB2), a highly conserved scaffold protein, is abnormally expressed and activated in patients with gastric cancer (GC). However, the genetic diversity of GAB2 in GC and its association with the clinical manifestations of patients are still unclear. Here, we explored the polymorphism of GAB2 rs2373115 in GC and its association with quality of life (QOL) and negative emotions of patients with GC after postoperative comprehensive care. A case-control study showed that the frequency of the GG genotype of GAB2 rs2373115 in the GC patients was higher than that in the healthy people, while the frequency of the TT + TG genotype was lower than that in the healthy people. Obvious distinctions were observed in the histological grade and TNM staging between the GG genotype and TT + TG genotype. In addition, SAS and SDS scores in the patients with GG genotype were higher than those in patients with TT + TG genotype, while the emotional function, cognitive function, dyspnea, fatigue, sleep disorder, and overall QOL in patients with GG genotype were lower than those in patients with TT + TG genotype. These results showed that GAB2 rs2373115 polymorphism was related to QOL and negative emotions in patients with GC after postoperative comprehensive care.

Molecular profile reveals immune-associated markers of medulloblastoma for different subtypes.

Medulloblastoma, a common pediatric malignant tumor, has been recognized to have four molecular subgroups [wingless (WNT), sonic hedgehog (SHH), group 3, group 4], which are defined by the characteristic gene transcriptomic and DNA methylomic profiles, and has distinct clinical features within each subgroup. The tumor immune microenvironment is integral in tumor initiation and progression and might be associated with therapeutic responses. However, to date, the immune infiltrative landscape of medulloblastoma has not yet been elucidated. Thus, we proposed MethylCIBERSORT to estimate the degree of immune cell infiltration and weighted correlation network analysis (WGCNA) to find modules of highly correlated genes. Synthesizing the hub genes in the protein–protein interaction (PPI) network and modules of the co-expression network, we identify three candidate biomarkers [GRB2-associated-binding protein 1 (GAB1), Abelson 1 (ABL1), and CXC motif chemokine receptor type 4 (CXCR4)] via the molecular profiles of medulloblastoma. Given this, we investigated the correlation between these three immune hub genes and immune checkpoint blockade response and the potential of drug prediction further. In addition, this study demonstrated a higher presence of endothelial cells and infiltrating immune cells in Group 3 tumor bulk. The above results will be conducive to better comprehending the immune-related pathogenesis and treatment of medulloblastoma.

Circ_0057452 sponges miR-7-5p to promote keloid progression through upregulating GAB1

ABSTRACT Increasing evidence has shown that circular RNAs (circRNAs) play critical roles in various diseases, including keloid. The purpose of this study was to investigate the role of circ_0057452 and related action mechanisms during the development of keloid. The expression levels of circ_0057452, microRNA-7-5p (miR-7-5p) and GRB2 associated binding protein 1 (GAB1) mRNA were determined by quantitative real-time PCR (qRT-PCR). Cell proliferation was evaluated using methylthiazolyldiphenyl-tetrazolium bromide (MTT) and 5-Ethynyl-2’-deoxyuridine (Edu) assays. Flow cytometry analysis was utilized to determine cell cycle distribution and cell apoptosis. Western blot assay was used to measure apoptosis-related, collagen synthesis-related, and GAB1 protein levels. Cell migration and invasion were detected by wound healing assay and transwell assay. The interaction between miR-7-5p and circ_0057452 or GAB1 was confirmed by dual-luciferase reporter, RNA pull-down, and RNA Immunoprecipitation (RIP) assays. Circ_0057452 and GAB1 were upregulated in keloid tissues and keloid fibroblasts (KFs), while miR-7-5p was downregulated. Circ_0057452 knockdown or miR-7-5p overexpression inhibited the proliferation, migration, invasion, and collagen synthesis and induced cell cycle arrest and apoptosis of KFs. MiR-7-5p was targeted by circ_0057452, and its inhibition overturned the effects of circ_0057452 knockdown. In addition, GAB1 was a target of miR-7-5p, and GAB1 upregulation abolished the role of miR-7-5p overexpression and circ_0057452 knockdown in KFs. Circ_0057452 regulated the expression of GAB1 by adsorbing miR-7-5p in KFs. Circ_0057452 knockdown suppressed keloid development by regulating miR-7-5p/GAB1 axis, which might provide a promising therapeutic target for keloid.

Her2/EGFR-PDGFR pathway aberrations associated with tamoxifen response in metastatic breast cancer patients

BackgroundMetastatic breast cancer (MBC) is a major health problem worldwide. Some patients improve on tamoxifen and others do not respond to treatment. Therefore, the aim of the current study is to assess genetic aberrations in the Her2/EGFR-PDGFR pathway associated with tamoxifen response in MBC patients.MethodsThis is a retrospective cohort study, including 157 hormone receptors positive, locally recurrent inoperable and/or MBC patients on tamoxifen treatment. Patients were categorized into 78 (49.7%) tamoxifen responders and 79 (50.3%) tamoxifen non-responder patients. Genetic aberrations of 84 genes involved in the Her2/EGFR-PDGFR pathway were assessed in the tumor tissue samples obtained from the patients using SA-Bioscience assay. The identified panel was correlated to patients’ response to treatment, to detect the differentially expressed genes in tamoxifen responders and non-responders.ResultsOne hundred twenty-three (78.3%) patients were estrogen receptor (ER) and progesterone receptor (PR) positive, 108 (68.8%) were ER only positive, and 78 (49.7%) were PR only positive. There were 56 genes overexpressed in the refractory group compared to responders. However, only five out of these 56 genes, Janus kinase 1 (JAK1), collagen type I alpha 1 (COL1A1), GRB2-associated binding protein 1 (GAB1), fibronectin-1 (FN1), and MAP kinase-interacting serine/threonine-protein kinase (MKNK1), showed statistical significance between the two groups. Patients with bone metastasis showed a better response to treatment compared to those with metastatic deposits in other sites such as visceral metastasis (P < 0.005).ConclusionsGenetic profiling using simple quantitative real-time polymerase chain reaction (qRT-PCR) protocols could be used to assess response to tamoxifen treatment in MBC patients. According to our data, a five-gene panel in the EGFR pathway (JAK1, COL1A1, GAB1, FN1 and MKNK1) could be used to categorize MBC patients into groups according to treatment response.

Gab1 Overexpression Alleviates Doxorubicin-Induced Cardiac Oxidative Stress, Inflammation, and Apoptosis Through PI3K/Akt Signaling Pathway.

Grb2-associated binding protein 1 (Gab1), an intracellular scaffolding adaptor, was involved in several cardiovascular diseases. However, the role of Gab1 in doxorubicin (DOX)-induced cardiotoxicity remains largely unknown. The present study investigated whether Gab1 protected against DOX-induced cardiotoxicity and the underlying mechanism. We overexpressed Gab1 in the hearts using an adeno-associated virus 9 system through tail vein injection. C57BL/6 mice were subjected to DOX (15 mg/kg/d, i.p.) to generate DOX-induced cardiotoxicity. Echocardiography, histological analysis, immunofluorescence and enzyme-linked immunosorbent assay (ELISA) kits, Western blotting, and quantitative real-time polymerase chain reaction (PCR) evaluated DOX-induced cardiotoxicity and the underlying mechanisms. Myocardial Gab1 protein and messenger RNA (mRNA) levels were markedly decreased in DOX-administered mice. Overexpression of Gab1 in myocardium significantly improved cardiac function and attenuated cardiac oxidative stress, inflammatory response, and apoptosis induced by DOX. Mechanistically, we found that PI3K/Akt signaling pathway was downregulated after DOX treatment, and Gab1 overexpression activated PI3K/Akt signaling pathway, whereas PI3K/Akt signaling pathway inhibition abolished the beneficial effect of Gab1 overexpression in the heart. Collectively, our results indicated that Gab1 is essential for cardioprotection against DOX-induced oxidative stress, inflammatory response, and apoptosis by mediating PI3K/Akt signaling pathway. And cardiac gene therapy with Gab1 provides a novel therapeutic strategy against DOX-induced cardiotoxicity.

Identification of Grb2-associated binding protein 3 expression to predict clinical outcomes and immunotherapeutic responses in lung adenocarcinoma.

With the deepening research on tumor microenvironment (TME), immunotherapy has been deemed to be one of the major breakthroughs for cancer therapy. Nevertheless, only some patients respond well to this treatment. It is vital to explore predictive biomarkers for clinical benefit of immunotherapy. Grb2-associated binding protein 3 (GAB3) exerts essential biological functions in ovarian cancer and colorectal cancer. The potential role of GAB3 in lung adenocarcinoma (LUAD) has not been fully elucidated. RNA-sequencing data, genetic mutation data, and matched clinical data were obtained from the cancer genome atlas (TCGA) databases, then underwent gene expression, prognosis, enrichment, TME, immune checkpoint blockade (ICB) response analyses utilizing R packages. The mRNA expression level of GAB3 was dramatically decreased in LUAD, and the prognostic analysis indicated that the patients with low GAB3 expression performed unsatisfactory clinical outcomes. In addition, differentially expressed genes (DEGs) and subsequent functional enrichment analysis demonstrated that GAB3 was primarily connected with T cell activation and immune response. Finally, GAB3 expression positively correlated with immune infiltrates and immune checkpoint genes, and therapeutic effect of ICB. In summary, our study comprehensively uncovers that GAB3 may function as a promising biomarker to predict clinical outcomes and immunotherapeutic responses in LUAD patients.

Targeting SNORA38B attenuates tumorigenesis and sensitizes immune checkpoint blockade in non-small cell lung cancer by remodeling the tumor microenvironment via regulation of GAB2/AKT/mTOR signaling pathway.

BackgroundNon-coding RNAs (ncRNAs), including small nucleolar RNAs (snoRNAs), are widely involved in the physiological and pathological processes of human beings. While up to date, although considerable progress has been achieved in ncRNA-related pathogenesis of non-small cell lung cancer (NSCLC), the underlying mechanisms and biological significance of snoRNAs in NSCLC still need to be further clarified.MethodsQuantitative real-time polymerase chain reaction or RNAscope was performed to verify the expression of Small Nucleolar RNA, H/ACA Box 38B (SNORA38B) in NSCLC cell lines or clinical samples. BALB/c nude mice xenograft model or C57BL/6J mice syngeneic tumor model were estimated to detect the effects of SNORA38B in tumor growth or tumor immune microenvironment in vivo. Cytometry by time of flight, enzyme-linked immunosorbent assay and flow cytometry assay were conducted to clarify the effects and mechanisms of SNORA38B-mediated tumor immunosuppressive microenvironment. The binding activity between SNORA38B and E2F transcription factor 1(E2F1) was detected by RNA immunoprecipitation and RNA pull-down assays. Then, bioinformatics analysis and chromatin immunoprecipitation were utilized to demonstrate the regulation of GRB2-associated-binding protein 2 (GAB2) by E2F1. Moreover, the combinatorial treatment of SNORA38B locked nucleic acid (LNA) and immune checkpoint blockade (ICB) was used to treat murine Lewis lung carcinoma-derived tumor burden C57BL/6J mice to clarify the effectiveness of targeting SNORA38B in NSCLC immunotherapy.ResultsSNORA38B was found highly expressed in NSCLC tissues and cell lines, and associated with worse prognosis. Further results showed that SNORA38B functioned as an oncogene via facilitating cell proliferation, migration, invasion, and inhibiting cell apoptosis in vitro and promoting tumorigenesis of NSCLC cells in vivo. SNORA38B could also recruit the CD4+FOXP3+ regulatory T cells by triggering tumor cells to secrete interleukin 10, which in turn reduced the infiltration of CD3+CD8+ T cells in NSCLC tumor microenvironment (TME), favoring tumor progression and poorer immune efficacy. Mechanistically, SNORA38B mainly distributed in the nucleus, and promoted NSCLC progression by regulating GAB2 transcription to activate protein kinase B (AKT)/mammalian target of rapamycin (mTOR) pathway through directly binding with E2F1. Moreover, we found that SNORA38B LNAs were able to ameliorate CD3+CD8+ T cell infiltration in TME, which sensitized NSCLC to the treatment of ICB.ConclusionsIn conclusion, our data demonstrated that SNORA38B functioned as an oncogene in NSCLC both in vitro and in vivo at least in part by regulating the GAB2/AKT/mTOR pathway via directly binding to E2F1. SNORA38B could also sensitize NSCLC to immunotherapy, which may be a critical therapeutic target for NSCLC.

Mechanism of METTL3-mediated m6A modification in depression-induced cognitive deficits.

Depressive disorder (DD) is associated with N6-methyladenosine (m6A) hypermethylation. This study sought to explore the molecular mechanism of Methyltransferase-like 3 (METTL3) in cognitive deficits of chronic unpredictable mild stress (CUMS)-treated rats and provide novel targets for DD treatment. A DD rat model was established via CUMS treatment. Cognitive deficits were assessed via body weighing and behavioral tests. METTL3, microRNA (miR)-221-3p, pri-miR-221, GRB2-associated binding protein 1 (Gab1) expressions in hippocampal tissues were detected via RT-qPCR and Western blotting. m6A, DiGeorge syndrome critical region gene 8 (DGCR8)-bound pri-miR-221 and pri-miR-221 m6A levels were measured. The binding relationship between miR-221-3p and Gab1 was testified by dual-luciferase and RNA pull-down assays. Rescue experiments were designed to confirm the role of miR-221-3p and Gab1. METTL3 was highly expressed in CUMS rats, and silencing METTL3 attenuated cognitive deficits of CUMS rats. METTL3-mediated m6A modification facilitated processing and maturation of pri-miR-221 via DGCR8 to upregulate miR-221-3p. miR-221-3p targeted Gab1. miR-221-3p overexpression or Gab1 downregulation reversed the role of silencing METTL3 in CUMS rats. Overall, METTL3-mediated m6A modification facilitated processing and maturation of pri-miR-221 to upregulate miR-221-3p and then inhibit Gab1, thereby aggravating cognitive deficits of CUMS rats.

The comparison of differentially expressed microRNAs in Bag-1 deficient and wild type MCF-7 breast cancer cells by small RNA sequencing

The multifunctional BAG-1 (Bcl-2 athanogene-1) protein promotes breast cancer survival through direct or indirect interaction partners. The number of the interacting partners determines its cellular role in different conditions. As well as interaction partner variability, the amount of BAG-1 protein in the cells could cause dramatic alterations. According to previous studies, while the transient silencing of Bag-1 enhanced drug-induced apoptosis, deletion of BAG-1 could induce stemness properties and Akt-mediated actin remodeling in MCF-7 breast cancer cells. Considering the heterogeneity of breast cancer and the variability of BAG-1 -mediated cell response, it has become essential to determine microRNA (miRNA) functions in breast cancer depending on Bag-1 expression level. This study aims to compare microRNA expression levels in wt and Bag-1 knockout (KO) MCF-7 breast cancer cells. hsa-miR-429 was selected as a potential miRNA in BAG-1KO MCF-7 cells because of the downregulation both in bioinformatics and validation qRT-PCR assay. According to predicted mRNA targets and functional enrichment analysis the ten hub proteins that are phosphatidylinositol- 4,5-biphosphate 3-kinase catalytic subunit alpha (PIK3CA), kinase insert domain receptor (KDR), GRB2 associated binding protein 1 (GAB1), Rac family small GTPase1 (RAC1), vascular endothelial growth factor A (VEGFA), Cbl proto-oncogene (CBL), syndecan 2 (SDC2), phospholipase C gamma 1 (PLCG1), E1A binding protein p300 (EP300), and CRK like proto-oncogene, adaptor protein (CRKL) were identified as targets of hsa-miR-429. The functional enrichment analysis showed that the most significant proteins were enriched in PI3K/Akt, focal adhesion, cytoskeleton regulation, proteoglycans in cancer, and Ras signaling pathways. It was determined that hsa-miR-429 targeted these pathways in Bag-1 deficient conditions and could be used as a potential therapeutic target in future studies.