FHA Domains Research Articles

The Role of AGGF1 in the Classification and Evaluating Prognosis of Adult Septic Patients: An Observational Study.

Angiogenic factor with G patch and FHA domains 1 (AGGF1) is a crucial angiogenic factor that is involved in a variety of diseases and in the regulation of inflammatory responses. However, its role in sepsis is poorly understood. We have investigated the role of AGGF1 in the classification and prognostic evaluation of adult septic patients in a clinical context. A total of 126 septic patients who visited the Emergency Department of Beijing Chao-Yang Hospital and 76 non-sepsis patients visiting the Physical Examination Center of Beijing Chao-Yang Hospital were enrolled. AGGF1 levels in plasma were detected by enzyme-linked immunosorbent assay. Spearman correlation analysis was used to determine correlations between plasma AGGF1 and Sequential Organ Failure Assessment (SOFA) score, Acute Pathology and Chronic Health Evaluation II (APACHE II) score, procalcitonin and lactate. We evaluated the classification significance of AGGF1 in sepsis using receiver operating characteristic (ROC) curves. We also assessed the predictive significance of AGGF1 for 28-day mortality in sepsis using ROC curves and Kaplan-Meier analyses. Plasma AGGF1 levels were higher in sepsis patients than in non-sepsis patients (P < 0.001). Among sepsis patients, plasma AGGF1 levels were higher in non-survivors than in survivors (P < 0.001). Increased plasma AGGF1 levels were positively correlated with SOFA score, APACHE II score, procalcitonin and lactate. Plasma AGGF1 levels could distinguish sepsis patients from non-sepsis patients (area under the curve [AUC] = 0.777). AGGF1 had a higher predictive value than SOFA score, APACHE II score, lactate, procalcitonin, and white blood cell count for 28-day mortality in patients with sepsis (AUC = 0.876). Furthermore, Kaplan-Meier analysis indicated that lower plasma AGGF1 levels were associated with lower 28-day mortality compared with higher plasma AGGF1 levels (log rank P < 0.001). AGGF1 is useful for the classification and evaluating prognosis of adult septic patients.

2nd Workshop in Microbial Molecular Biology

2nd Workshop in Microbial Molecular Biology

Endothelial G protein stimulatory α-subunit is a critical regulator of post-ischemic angiogenesis.

Post-ischemic angiogenesis is a vital pathophysiological process in diseases such as peripheral arterial disease (PAD), heart ischemia, and diabetic retinopathy. The molecular mechanisms of post-ischemic angiogenesis are complicated and not fully elucidated. The G protein stimulatory alpha subunit (Gsα) is essential for hormone-stimulated cyclic adenosine monophosphate (cAMP) production and is an important regulator for many physiological processes. In the present study, we investigated the role of endothelial Gsα in post-ischemic angiogenesis by generating adult mice with endothelial-specific Gsα deficiency (GsαECKO). GsαECKO mice had impaired blood flow recovery after hind limb ischemic injury, and reduced neovascularization in allograft transplanted tumors. Mechanically, Gsα could regulate the expression of angiogenic factor with G patch and FHA domains 1 (AGGF1) through cAMP/CREB pathway. AGGF1 plays a key role in angiogenesis and regulates endothelial cell proliferation as well as migration. Knockdown of CREB or mutation of the CRE site on the AGGF1 promoter led to reduced AGGF1 promoter activity. In addition, knockdown of AGGF1 reduced the proangiogenic effect of Gsα in endothelial cells, and overexpression of AGGF1 reversed the impaired angiogenesis in GsαECKO mice in vivo. The finding may prove useful in designing new therapeutic targets for treatments of post-ischemic angiogenesis-related diseases.

The architecture of kinesin-3 KLP-6 reveals a multilevel-lockdown mechanism for autoinhibition

Autoinhibition of kinesin-3 ensures the proper spatiotemporal control of the motor activity for intracellular transport, but the underlying mechanism remains elusive. Here, we determine the full-length structure of kinesin-3 KLP-6 in a compact self-folded state. Unexpectedly, all the internal coiled-coil segments and domains in KLP-6 cooperate to successively lock down the neck and motor domains. The first coiled-coil segment is melted into several short helices that work with the motor domain to restrain the entire neck domain. The second coiled-coil segment associates with its neighboring FHA and MBS domains and integrates with the tail MATH domain to form a supramodule that synergistically wraps around the motor domain to trap the nucleotide and hinder the microtubule binding. This multilevel-lockdown mechanism for autoinhibition could be applicable to other kinesin-3 motors.

Identification and characterization of a special type of subnuclear structure: AGGF1-coated paraspeckles.

AGGF1 is an angiogenic factor with G-Patch and FHA domains 1 described by our group. Gain-of-function mutations in AGGF1 cause Klippel-Trenaunay syndrome, whereas somatic loss-of-function mutations cause cancer. Paraspeckles are small membraneless subnuclear structures with a diameter of 0.5-1μm, and composed of lncRNA NEAT1 as the scaffold and three core RNA-binding proteins NONO, PSPC1, and PSF. Here, we show that AGGF1 is a key regulatory and structural component of paraspeckles that induces paraspeckle formation, forms an outside rim of paraspeckles, wraps around the NONO/PSF/PSPC1/NEAT1 core, and regulates the size and number of paraspeckles. AGGF1-paraspeckles are larger (>1μm) than conventional paraspeckles. RNA-FISH in combination with immunostaining shows that AGGF1, NONO, and NEAT1_2 co-localize in 20.58% of NEAT1_2-positive paraspeckles. Mechanistically, AGGF1 interacts with NONO, PSF, and HNRNPK, and upregulates NEAT1_2, a longer, 23kb NEAT1 transcript with a key role in regulation of paraspeckle size and number. RNA-immunoprecipitation shows that AGGF1 interacts with NEAT1, which may be another possible mechanism underlying the formation of AGGF1-paraspeckles. NEAT1_2knockdown reduces the number and size of AGGF1-paraspeckles. Functionally, AGGF1 regulates alternative RNA splicing as it decreases the exon skipping/inclusion ratio in a CD44model. AGGF1 is also localized in some nuclear foci without NEAT1 or NONO, suggesting that AGGF1 is an important liquid-liquid phase separation (LLPS) driver for other types of AGGF1-positive nuclear condensates (referred to as AGGF1-bodies). Our results identify a special type of AGGF1-coated paraspeckles and provide important insights into the formation, structure, and function of paraspeckles.

Neuroprotective effect of AGGF1 against isoflurane-induced cognitive dysfunction in aged rats through activating the PI3K/AKT signaling pathways.

This study aimed to evaluate and identify the value and explore the mechanisms of Angiogenic Factor with G-patch and FHA domains 1 (AGGF1) in postoperative cognitive dysfunction (POCD). Rats were separated into four different groups, namely sham, isoflurane, isoflurane + recombinant human Aggf1 (rh-Aggf1) (5 μg kg-1), and isoflurane + rh-Aggf1 (10 μg kg-1). qPCR and western blot assays were applied to detect the correlation between the expression of AGGF1 and isoflurane administration. Then, the Morris water maze (MWM) test was applied to evaluate the effect of AGGF1 on improving the POCD rats. Subsequently, TUNEL assay was applied and the cell apoptosis-related proteins were tested to reveal the anti-apoptotic effect of AGGF1 in POCD rats. Furthermore, the mRNA and protein levels of TNF-α, IL-6, and IL-1β were also detected by qPCR and ELISA to verify the anti-inflammatory effects of AGGF1 on POCD rats. Besides, the protein expression levels of PI3K, Akt, and NF-κB in each group were examined by western blot. In this study, the results revealed that isoflurane induced a decrease in AGGF1 expression in the hippocampus of aged rats. In addition, exogenous AGGF1 attenuated POCD in aged rats. Meanwhile, exogenous AGGF1 had anti-apoptotic and anti-inflammatory effects in POCD rats. Further research indicated that AGGF1 activated the PI3K/Akt pathway. AGGF1 has neuroprotective effect against isoflurane-induced cognitive dysfunction in aged rats via activating the PI3K/AKT signaling pathways.

Expression and Clinical Significance of the NCAPH, AGGF1, and FOXC2 Proteins in Serous Ovarian Cancer.

PurposeRecurrence and metastasis are the most common causes of high mortality rates in patients with serous ovarian cancer (SOC). Non-structural maintenance of chromosomes (non-SMC) condensin I complex subunit H (NCAPH) is a newly identified essential oncoprotein whose function in SOC pathogenesis has not been reported yet. Angiogenic factor with G patch and FHA domains 1 (AGGF1) is an effective promoter of angiogenesis in humans, leading to cancer cell infiltration and progression. Forkhead box C2 (FOXC2) plays a pivotal role in epithelial-to-mesenchymal transition (EMT). The present study analyzed the correlations among the expressions of these three proteins and their relationships with the clinicopathological characteristics and survival of patients with SOC.Patients and MethodsThe expressions of NCAPH, AGGF1, and FOXC2 were detected by the immunohistochemical examination of 153 SOC tissue samples and 30 serous ovarian cystadenoma tissue samples. Clinicopathologic and follow-up data of the patients were collected.ResultsThe expressions of NCAPH, AGGF1, and FOXC2 were remarkably higher in the SOC tissue samples than in the serous ovarian cystadenoma tissue samples. The protein expressions were positively correlated with the histological tumor grade, the International Federation of Gynecology and Obstetrics (FIGO) stage, lymph node metastasis, and intraperitoneal implantation, but were negatively correlated with the overall survival (OS). Moreover, multivariate analysis showed that the NCAPH, AGGF1, and FOXC2 expressions, FIGO stage, and histological tumor grade were independent adverse prognostic factors for OS in patients with SOC.ConclusionThe results of this study show that the expressions of NCAPH, AGGF1, and FOXC2 are promising biomarkers and possible therapeutic targets in patients with SOC.

The Relationship Between UBE2C and AGGF1 Overexpression and Tumor Angiogenesis in Non-Small Cell Lung Cancer.

BackgroundTumor infiltration and metastasis are the leading causes of death for patients with tumors. Angiogenesis is a prerequisite for tumor growth and metastasis. Angiogenic factor with G patch and FHA domains 1 (AGGF1) is an angiogenic factor, whereas ubiquitin-conjugating enzyme E2C (UBE2C) functions in protein ubiquitination. Microvessel density (MVD) is the most common indicator of tumor microvessels, and vasculogenic mimicry (VM) facilitates blood supply to tumors. This study explored UBE2C and AGGF1 expression in non-small cell lung cancer (NSCLC) and their relationship with angiogenesis and prognosis to identify biological factors that might predict NSCLC infiltration, metastasis, and prognosis.MethodsThe specimens and clinical pathological data of patients with NSCLC confirmed by pathology after surgical resection between January 2013 and December 2015 were collected. UBE2C and AGGF1 expression, as well as microvessel formation and VM in NSCLC, was observed using immunohistochemistry. The relationships between UBE2C, AGGF1, MVD, VM, and clinical pathological parameters and their relationships with overall survival (OS) and disease-free survival (DFS) were analyzed.ResultsUBE2C and AGGF1 levels in NSCLC tissues were significantly higher than those in corresponding normal tissues (57.1% vs 15.6 and 59.7% vs 25.3%, respectively; P < 0.05). UBE2C, AGGF1, MVD, and VM were positively correlated with each other (P < 0.05) and were all related to tumor size, lymph node metastasis, and tumor-node-metastasis stage (P < 0.05). Kaplan–Meier analysis showed that patient OS and DFS in the UBE2C, AGGF1, VM-positive, and high-MVD groups were reduced (all P < 0.001). Univariate and multivariate analyses showed that UBE2C, AGGF1, VM, and MVD were independent risk factors for NSCLC prognosis.ConclusionUBE2C and AGGF1 overexpression is associated with angiogenesis and poor prognosis and may be important for predicting NSCLC invasion, metastasis, and prognosis.

CHFR‑mediated epithelial‑to‑mesenchymal transition promotes metastasis in human breast cancer cells.

Checkpoint with FHA and RING finger domains (CHFR) is a G2 phase/mitosis checkpoint. Several studies have reported that CHFR is downregulated in multiple cancer types and serves a tumor suppressor role. However, the biological function of CHFR in breast cancer (BRCA), particularly regarding metastasis, are yet to be elucidated. In the present study, it was revealed that CHFR is upregulated in BRCA compared with normal tissues, according to The Cancer Genome Atlas database. In addition, subgroup analysis of BRCA revealed that CHFR was upregulated in both human epidermal growth factor receptor2‑positive and triple‑negative BRCA. Meanwhile, patients with high expression levels of CHFR exhibited poorer overall survival rates. Furthermore, the present data revealed that the overexpression of CHFR in SKBR3 cells resulted in enhanced cell migration and invasiveness, and also significantly upregulated mesenchymal markers, such as N‑cadherin, vimentin, transcription factor Slug and tight junction protein claudin‑1. Furthermore, knockdown of CHFR in MDA‑MB‑231 cells significantly inhibited cell migration and invasiveness, and also downregulated mesenchymal markers, such as N‑cadherin, vimentin and tight junction protein claudin‑1. In conclusion, the current results indicated that CHFR expression was associated with cell metastasis in BRCA by mediating epithelial‑to‑mesenchymal transition.

Structure of the Mycobacterium tuberculosis cPknF and conformational changes induced in forkhead-associated regulatory domains.

Mycobacterium tuberculosis (Mtb) has 11 Serine-Threonine Protein Kinases (STPK) that control numerous physiological processes, including cell growth, cell division, metabolic flow, and transcription. PknF is one of the 11 Mtb STPKs that has, among other substrates, two FHA domains (FHA-1 and FHA-2) of the ATP-Binding Cassette (ABC) transporter Rv1747. Phosphorylation in T152 and T210 located in a non-structured linker that connects Rv1747 FHA domains is considerate to be the regulatory mechanism of the transporter. In this work, we resolved the three-dimensional structure of the PknF catalytic domain (cPknF) in complex with the human kinase inhibitor IKK16. cPknF is conserved when compared to other STPKs but shows specific residues in the binding site where the inhibitor is positioned. In addition, using Small Angle X-Ray Scattering analysis we monitored the behavior of the wild type and three FHA-phosphomimetic mutants in solution, and measured the cPknF affinity for these domains. The kinase showed higher affinity for the non-phosphorylated wild type domain and preference for phosphorylation of T152 inducing the rapprochement of the domains and significant structural changes. The results shed some light on the process of regulating the transporter's activity by phosphorylation and arises important questions about evolution and importance of this mechanism for the bacillus.

CircSFMBT2 facilitates vascular smooth muscle cell proliferation by targeting miR-331-3p/HDAC5

ObjectiveTo investigate the functional role of circSFMBT2 in vascular smooth muscle cell (VSMC) proliferation and migration and the underlying molecular mechanism. MethodsThe circSFMBT2 levels in neointimal tissue and platelet derived growth factor-BB (PDGF-BB)-treated VSMCs were detected by qRT-PCR. The role of circSFMBT2 in VSMC proliferation, migration and cell cycle distribution was assessed by MTT assay, transwell assay, wound healing assay and flow cytometry. The protein expression of contractile markers was evaluated by western blot. In vitro luciferase reporter assay, RNA pull-down assay, ChIP and coimmunoprecipitation (CoIP) were performed to explore the effects of circSFMBT2 on the downstream signaling pathway. ResultsWe found that circSFMBT2 was markedly increased in neointimal tissue relative to normal tissue and PDGF-BB-treated VSMCs relative to control VSMCs. The knockdown of circSFMBT2 by siRNA significantly inhibited the proliferation and migration of VSMCs. Interestingly, circSFMBT2 knockdown enhanced the expression of contractile marker proteins including SM22α, SM myosin heavy chain (SMMHC) and calponin. Further data demonstrated that circSFMBT2 interacted with miR-331-3p as a competing endogenous RNA and up-regulated the expression of histone deacetylase 5 (HDAC5), thereby regulating the level of angiogenic factor with G patch and FHA domains (Aggf1). ConclusionThese results revealed that circSFMBT2 plays a vital role in VSMC proliferation and migration through the miR-331/HDAC5/Aggf1 axis, and suggest a novel target for treating proliferative vascular diseases.

Expression of angiogenic factor with G patch and FHA domains 1 (AGGF1) in placenta from patients with preeclampsia.

Preeclampsia (PE) is a major contributor to maternal and foetal morbidity and mortality worldwide. It manifests as high blood pressure and proteinuria in women at more than 20 weeks of gestation. Abnormal levels of anti- and pro-angiogenesis factors are known to be associated with PE. In the present study, we aimed to determine the localisation of angiogenic factor with G patch and FHA domains 1 (AGGF1) in the placenta and to compare the expression levels of AGGF1 in the third-trimester placentas of preeclamptic and normotensive pregnancies. Placental tissue samples were collected from women with PE (n = 28) and without PE (n = 28). The normotensive controls without PE were matched for gestational age at delivery with the patients with PE. The expression levels of AGGF1 in the placental tissues were evaluated using immunohistochemistry, quantitative reverse transcription polymerase chain reaction and Western blot. The immunoexpression of AGGF1 was localised in the syncytiotrophoblast tissue. Notable, the mRNA and protein expression levels of AGGF1 were decreased in preeclamptic placentas as compared with the normotensive control group (P < 0.05). Our results suggest that the decreased AGGF1 in preeclamptic placentas may be related to the pathogenesis of preeclampsia.

Angiogenic factor with G patch and FHA domains 1 protects retinal vascular endothelial cells under hyperoxia by inhibiting autophagy.

Angiogenic factor with G patch and FHA domains 1 (AGGF1) has strong proangiogenic effects on embryonic vascular development and angiogenesis in disease; however, its role in retinopathy has not been elucidated. Retinopathy of prematurity is a serious retinal disorder of premature infants, which is caused by the arrest of immature retinal vascular growth under hyperoxia. This study aims to investigate the effects of AGGF1 on retinal vascular endothelial cells under hyperoxia and the association with autophagy by using rhesus macaque choroid-retinal endothelial (RF/6A) cells. Western blot analysis and immunofluorescence staining were used to detect the expression of AGGF1 in RF/6A cells. Cell Counting Kit-8, flow cytometry, and transwell and matrigel assays were applied to detect the vitality, apoptosis, migration, and tube formation of RF/6A cells, respectively. Western blot analysis was then used to detect the expression of autophagy markers LC3 and Beclin-1, and mCherry-GFP-LC3 adenovirus was used to detect autophagy flux in RF/6A cells. Under hyperoxia, the expression of AGGF1 in RF/6A cells decreased compared with the control. Cell vitality, migration, and tube formation decreased, and apoptosis of RF/6A cells increased under hyperoxia, and these effects of hyperoxia were attenuated by AGGF1. The protein expressions of LC3 and Beclin-1 increased in RF/6A cells and autophagy flux enhanced under hyperoxia. AGGF1 reduced the expression of LC3 and Beclin-1 as well as the autophagy flux stimulated by hyperoxia. The results clearly showed that exogenous AGGF1 can protect retinal vascular endothelial cells and promote angiogenesis under hyperoxia, in which the expression of AGGF1 was inhibited. Inhibition of autophagy by AGGF1 may be one of the mechanisms involved.

Vaccarin Regulates Diabetic Chronic Wound Healing through FOXP2/AGGF1 Pathways.

Background: Diabetes mellitus is a growing global health issue nearly across the world. Diabetic patients who are prone to develop diabetes-related complications often exhibit progressive neuropathy (painless and sensory loss). It is usual for small wounds to progress to ulceration, which especially worsens with peripheral arterial disease and in the presence of anaerobic bacteria, culminating into gangrene. In our study, vaccarin (VAC), the main active monomer extracted from Chinese herb vaccariae semen, is proven to have a role in promoting diabetic chronic wound healing through a cytoprotective role under high glucose conditions. Materials and methods: We constructed a pressure ulcer on both VAC-treated and control mice based on a type 1 diabetes (T1DM) model. The wound healing index was evaluated by an experimental wound assessment tool (EWAT). We also determined the effect of VAC on the proliferation and cell migration of human microvascular endothelial cells (HMEC-1) by a cell counting kit (CCK-8), a scratch and transwell assay. Results: The results demonstrated that VAC could promote the proliferation and migration of high glucose-stimulated HMEC-1 cells, which depend on the activation of FOXP2/AGGF1. Activation of the angiogenic factor with G patch and FHA domains 1 (AGGF1) caused enhanced phosphorylation of serine/threonine kinase (Akt) and extracellular regulated protein kinases (Erk1/2). By silencing the expression of forkhead box p2 (FOXP2) protein by siRNA, both mRNA and protein expression of AGGF1 were downregulated, leading to a decreased proliferation and migration of HMEC-1 cells. In addition, a diabetic chronic wound model in vivo unveiled that VAC had a positive effect on chronic wound healing, which involved the activation of the above-mentioned pathways. Conclusions: In summary, our study found that VAC promoted chronic wound healing in T1DM mice by activating the FOXP2/AGGF1 pathway, indicating that VAC may be a promising candidate for the treatment of the chronic wounds of diabetic patients.

Angiogenic Factor with G Patch and FHA Domains 1 (AGGF1) Acts as Diagnostic Biomarker and Adverse Prognostic Factor of Hepatocellular Carcinoma (HCC): Evidence from Bioinformatic Analysis

BackgroundAngiogenic factor with G patch and FHA domains 1 (AGGF1) is a novel identified initiator of angiogenesis through promoting the proliferation of endothelial cells. The continuous angiogenesis plays a key role in the growth, invasion, and metastasis of hepatocellular carcinoma (HCC), while the diagnostic and prognostic roles of AGGF1 for HCC need to be further studied.Material/MethodsThe mRNA sequencing datasets and clinical features of HCC patients were extracted from The Cancer Genome Atlas database. The relationship between clinical features and AGGF1 expression was analyzed by Wilcoxon test. Further validation explorations were carried out using online database Oncomine. The diagnostic receiver operating characteristic curves of AGGF1 and alpha-fetoprotein were compared to examine the diagnostic efficacy of AGGF1. Survival analysis and Gene Set Enrichment Analysis were performed to explore the prediction value and potential mechanism of AGGF1 dysregulation in HCC.ResultsComprehensive overexpression of AGGF1 was observed in HCC, correlating with poor overall survival. Upregulated level of AGGF1 was statistically associated with poor differentiated histological grade, advanced cancer stage and T classification. AGGF1 was a more effective diagnostic marker than alpha-fetoprotein in HCC. Several important pathways related to HCC including pathway in cancer and P53 signaling pathway were differentially enriched in the high AGGF1 expression phenotype.ConclusionsAGGF1 was a potential diagnostic and prognostic marker for poor clinical outcomes in HCC patients. Moreover, vital pathways regulated by AGGF1 in HCC may include regulation of autophagy, Wnt signaling pathway, pathway in cancer, cell cycle, and P53 signaling pathway.

Elevated expression of AGGF1 predicts poor prognosis and promotes the metastasis of colorectal cancer

BackgroundAngiogenic factor with G-patch and FHA domains 1 (AGGF1) can promote angiogenesis and increasing evidence has highlighted the important roles of AGGF1 in tumorigenesis. However, the differential expression as well as the biological functions of AGGF1 in colorectal cancer (CRC) remain to be established. The purpose of the present study is therefore to identify the effect of AGGF1 on prognosis and metastasis in CRC patients.MethodsThe expression level of AGGF1 in CRC was examined by qPCR, western blot and immunohistochemistry in a tissue microarray containing 236 CRC specimens and paired normal mucosae. And the effect of AGGF1 on CRC cell malignance was investigated in our established stable AGGF1 upregulated and knockdown CRC cell lines.ResultsThe expression level of AGGF1 in CRC tissue was not significantly different to that in adjacent normal mucosa at the mRNA level. However, at the protein level, AGGF1 expression in CRC tissues was significantly higher than in paired normal mucosa, which showed a clear association with TNM stage, AJCC stage, vascular invasion, and differentiation. Further, we revealed an apparent correlation between AGGF1 expression and poorer disease-free survival and overall survival of CRC patients. In addition, we discovered that AGGF1 significantly promoted CRC cell wound healing, migration, and invasion in vitro and distant metastasis in vivo.ConclusionsOur study demonstrates the aberrant overexpression of AGGF1 in CRC and provides a basis on which to explore the application of AGGF1 as a potential therapeutic target for CRC patients, especially for CRC patients with distant metastasis.

2-Methoxyestradiol promotes radiosensitivity of esophageal squamous cell carcinoma by suppressing hypoxia-inducible factor-1α expression.

Acquired radioresistance remains the primary obstacle to improving the survival of esophageal cancer (EC). Related hypoxia factors play crucial roles in radioresistance such as hypoxia-inducible factor 1α (HIF-1α), vascular endothelial growth factor (VEGF), and angiogenic factor with G-patch and FHA domains 1 (AGGF1). The aim of this study was to investigate the effect of 2-methoxyestradiol (2ME2) on the radiosensitivity of EC. The expression of HIF-1α, VEGF, and AGGF1 in 70 EC patients was detected using immunohistochemistry. The relation between the expression levels of the above genes with clinicopathologic characteristics was explored as well. Human ECA-109 cells were subjected to hypoxia and/or radiation in the presence or absence of 2ME2. Subsequently, cell growth, colony formation, and apoptosis were evaluated. Moreover, the mRNA and protein expression levels were determined using quantitative Real Time-Polymerase Chain Reaction (qRT-PCR) and Western blotting, respectively. HIF-1α, VEGF, and AGGF1 were significantly in EC tissues and were associated with cancer aggressiveness. 2ME2 treatment significantly increased the radiosensitivity of ESCC cells in a dose-dependent manner, which was related to the inhibited expression of HIF-1α. Subsequent immunohistochemical staining results showed that the VEGF expression was positively correlated with the HIF-1α expression in EC. High levels of HIF-1α, VEGF, and AGGF1 in EC are indicators of poor prognosis in patients treated with radiotherapy, which can offer new ideas and methods for the treatment of ESCC. In addition, 2ME2 inhibits the expression of HIF-1α and confers radiosensitivity in ECA-109 cells. Our findings suggest that 2ME2 has the potential to be applied as an adjuvant treatment with radiotherapy for EC.

NBS1 promotes the endonuclease activity of the MRE11-RAD50 complex by sensing CtIP phosphorylation.

DNA end resection initiates DNA double-strand break repair by homologous recombination. MRE11-RAD50-NBS1 and phosphorylated CtIP perform the first resection step via MRE11-catalyzed endonucleolytic DNA cleavage. Human NBS1, more than its homologue Xrs2 in Saccharomyces cerevisiae, is crucial for this process, highlighting complex mechanisms that regulate the MRE11 nuclease in higher eukaryotes. Using a reconstituted system, we show here that NBS1, through its FHA and BRCT domains, functions as a sensor of CtIP phosphorylation. NBS1 then activates the MRE11-RAD50 nuclease through direct physical interactions with MRE11. In the absence of NBS1, MRE11-RAD50 exhibits a weaker nuclease activity, which requires CtIP but not strictly its phosphorylation. This identifies at least two mechanisms by which CtIP augments MRE11: a phosphorylation-dependent mode through NBS1 and a phosphorylation-independent mode without NBS1. In support, we show that limited DNA end resection occurs invivo in the absence of the FHA and BRCT domains of NBS1. Collectively, our data suggest that NBS1 restricts the MRE11-RAD50 nuclease to S-G2 phase when CtIP is extensively phosphorylated. This defines mechanisms that regulate the MRE11 nuclease in DNA metabolism.

The E3 ubiquitin ligase Pellino2 mediates priming of the NLRP3 inflammasome

The NLRP3 inflammasome has an important function in inflammation by promoting the processing of pro-IL-1β and pro-IL-18 to their mature bioactive forms, and by inducing cell death via pyroptosis. Here we show a critical function of the E3 ubiquitin ligase Pellino2 in facilitating activation of the NLRP3 inflammasome. Pellino2-deficient mice and myeloid cells have impaired activation of NLRP3 in response to toll-like receptor priming, NLRP3 stimuli and bacterial challenge. These functions of Pellino2 in the NLRP3 pathway are dependent on Pellino2 FHA and RING-like domains, with Pellino2 promoting the ubiquitination of NLRP3 during the priming phase of activation. We also identify a negative function of IRAK1 in the NLRP3 inflammasome, and describe a counter-regulatory relationship between IRAK1 and Pellino2. Our findings reveal a Pellino2-mediated regulatory signaling system that controls activation of the NLRP3 inflammasome.

Angiotensin II increases angiogenesis by NF‐κB–mediated transcriptional activation of angiogenic factor AGGF1

Angiogenic factor with G-patch and FHA domains 1 (AGGF1) is involved in vascular development, angiogenesis, specification of hemangioblasts, and differentiation of veins. When mutated, however, it causes Klippel-Trenaunay syndrome, a vascular disorder. In this study, we show that angiotensin II (AngII)-the major effector of the renin-angiotensin system and one of the most important regulators of the cardiovascular system-induces the expression of AGGF1 through NF-κB, and that AGGF1 plays a key role in AngII-induced angiogenesis. AngII significantly up-regulated the levels of AGGF1 mRNA and protein in HUVECs at concentrations of 10-40 μg/ml but not >60 μg/ml. AngII type 1 receptor (AT1R) inhibitor losartan inhibited AngII-induced up-regulation of AGGF1, whereas AT2R inhibitor PD123319 further increased AngII-induced up-regulation of AGGF1. Up-regulation of AGGF1 by AngII was blocked by NF-κB inhibitors, and p65 binds directly to a binding site at the promoter/regulatory region of AGGF1 and transcriptionally activates AGGF1 expression. AngII-induced endothelial tube formation was blocked by small interfering RNAs (siRNAs) for RELA (RELA proto-oncogene, NF-κB subunit)/p65 or AGGF1, and the effect of RELA siRNA was rescued by AGGF1. AngII-induced angiogenesis from aortic rings was severely impaired in Aggf1+/- mice, and the effect was restored by AGGF1. These data suggest that AngII acts as a critical regulator of AGGF1 expression through NF-κB, and that AGGF1 plays a key role in AngII-induced angiogenesis.-Si, W., Xie, W., Deng, W., Xiao, Y., Karnik, S. S., Xu, C., Chen, Q., Wang, Q. K. Angiotensin II increases angiogenesis by NF-κB-mediated transcriptional activation of angiogenic factor AGGF1.