Activation of the PERK/eIF2α axis is a pivotal prerequisite of taxanes to cancer cell apoptosis and renders synergism to overcome paclitaxel resistance in breast cancer cells.

BACKGROUND: Microtubule polymerization is usually considered as the upstream of apoptotic cell death induced by taxanes, but recently published studies provide more insights into the mechanisms responsible for the antineoplastic effect of taxanes. In this study, we figure out the role of the stress-related PERK/eIF2α axis in tumor cell death upon taxane treatment along with paclitaxel resistance. METHODS: Utilizing immunoblot assay, the activation status of PERK-eIF2α signaling was detected in a panel of cancer cell lines after the treatment of taxanes. The causal role of PERK-eIF2α signaling in the cancer cell apoptosis induced by taxanes was examined via pharmacological and genetic inhibitions of PERK. The relationship between microtubule polymerization and PERK-eIF2α activation was explored by immunofluorescent and immunoblotting assays. Eventaually, the combined therapeutic effect of paclitaxel (PTX) and CCT020312, a PERK agonist, was investigated in PTX-resistant breast cancer cells in vitro and in vivo. RESULTS: PERK-eIF2α axis was dramatically activated by taxanes in several cancer cell types. Pharmacological or genetic inhibition of PERK efficiently impaired taxane-induced apoptotic cell death, independent of the cellular microtubule polymerization status. Moreover, PTX was able to activate the PERK/eIF2α axis in a very low concentration without triggering microtubule polymerization. In PTX-resistant breast cancer cells, the PERK/eIF2α axis was attenuated in comparison with the PTX-sensitive counterparts. Reactivation of the PERK/eIF2α axis in the PTX-resistant breast cancer cells with PERK agonist sensitized them to PTX in vitro. Combination treatment of the xenografted PTX-resistant breast tumors with PERK agonist and PTX validated the synergic effect of PTX and PERK activation in vivo. CONCLUSION: Activation of the PERK/eIF2α axis is a pivotal prerequisite of taxanes to initiate cancer cell apoptosis, which is independent of the well-known microtubule polymerization-dependent manner. Simultaneous activation of PERK-eIF2α signaling would be a promising therapeutic strategy to overcome PTX resistance in breast cancer or other cancers.

Erk5 functions in modulation of zebrafish intestinal permeability.

The intestine of zebrafish consists of mucosa, muscularis and serosa. Intestinal epithelial cells (IECs) act as a physical and biochemical barrier to protect against invasion by external commensal bacteria. Cell junction is one of the crucial basis of the barrier function. When cell junctions were disrupted, intestinal permeability would be naturally impeded. Extracellular signal-regulated kinase 5 (ERK5), belonging to the Mitogen-activated protein kinase (MAPK) family, is involved in the normal physiological development of the cardiovascular system and nervous system. But the role of erk5 in intestinal morphogenesis and intestinal function is yet to know. Here, we showed that knockout of the erk5 in zebrafish larvae resulted in intestinal wall hypoplasia, including the thinned intestinal wall, reduced intestinal folds, and disrupted cell junctions. In addition, the intestinal permeability assay demonstrated that knockout of erk5 resulted in increased intestinal permeability. All of these showed that erk5 plays an essential role in the maintenance of intestinal barrier function. Thus, our data indicate that erk5 is a critical effector in intestinal morphogenesis and intestinal function, and dysfunction of erk5 would lead to intestinal diseases.

Vaxign2: the second generation of the first Web-based vaccine design program using reverse vaccinology and machine learning.

Vaccination is one of the most significant inventions in medicine. Reverse vaccinology (RV) is a state-of-the-art technique to predict vaccine candidates from pathogen's genome(s). To promote vaccine development, we updated Vaxign2, the first web-based vaccine design program using reverse vaccinology with machine learning. Vaxign2 is a comprehensive web server for rational vaccine design, consisting of predictive and computational workflow components. The predictive part includes the original Vaxign filtering-based method and a new machine learning-based method, Vaxign-ML. The benchmarking results using a validation dataset showed that Vaxign-ML had superior prediction performance compared to other RV tools. Besides the prediction component, Vaxign2 implemented various post-prediction analyses to significantly enhance users' capability to refine the prediction results based on different vaccine design rationales and considerably reduce user time to analyze the Vaxign/Vaxign-ML prediction results. Users provide proteome sequences as input data, select candidates based on Vaxign outputs and Vaxign-ML scores, and perform post-prediction analysis. Vaxign2 also includes precomputed results from approximately 1 million proteins in 398 proteomes of 36 pathogens. As a demonstration, Vaxign2 was used to effectively analyse SARS-CoV-2, the coronavirus causing COVID-19. The comprehensive framework of Vaxign2 can support better and more rational vaccine design. Vaxign2 is publicly accessible at http://www.violinet.org/vaxign2.

Development of full-length cell-culture infectious clone and subgenomic replicon for a genotype 3a isolate of hepatitis C virus.

Hepatitis C virus (HCV) genotype 3 is widely distributed, and genotype 3-infected patients achieve a lower cure rate in direct-acting antiviral (DAA) therapy and are associated with a higher risk of hepatic steatosis than patients with other genotypes. Thus, the study of the virology and pathogenesis of genotype 3 HCV is increasingly relevant. Here, we developed a full-length infectious clone and a subgenomic replicon for the genotype 3a isolate, CH3a. From an infected serum, we constructed a full-length CH3a clone, however, it was nonviable in Huh7.5.1 cells. Next, we systematically adapted several intergenotypic recombinants containing Core-NS2 and 5'UTR-NS5A from CH3a, and other sequences from a replication-competent genotype 2 a clone JFH1. Adaptive mutations were identified, of which several combinations facilitated the replication of CH3a-JFH1 recombinants; however, they failed to adapt to the full-length CH3a and the recombinants containing CH3a NS5B. Thus, we attempted to separately adapt CH3a NS5B-3'UTR by constructing an intragenotypic recombinant using 5'UTR-NS5A from an infectious genotype 3a clone, DBN3acc, from which L3004P/M in NS5B and a deletion of 11 nucleotides (Δ11nt) downstream of the polyU/UC tract of the 3'UTR were identified and demonstrated to efficiently improve virus production. Finally, we combined functional 5'UTR-NS5A and NS5B-3'UTR sequences that carried the selected mutations to generate full-length CH3a with 26 or 27 substitutions (CH3acc), and both revealed efficient replication and virus spread in transfected and infected cells, releasing HCV of 104.2 f.f.u. ml-1. CH3acc was inhibited by DAAs targeting NS3/4A, NS5A and NS5B in a dose-dependent manner. The selected mutations permitted the development of subgenomic replicon CH3a-SGRep, by which L3004P, L3004M and Δ11nt were proven, together with a single-cycle virus production assay, to facilitate virus assembly, release, and RNA replication. CH3acc clones and CH3a-SGRep replicon provide new tools for the study of HCV genotype 3.

Identification of STXBP6-IRF1 positive feedback loop in regulation of PD-L1 in cancer.

The clinical success of immune checkpoint blockade against diverse human cancers highlights the critical importance of insightful understanding into mechanisms underlying PD-L1 regulation. IFN-γ released by intratumoral lymphocytes regulates PD-L1 expression in tumor cells through JAK-STAT-IRF1 pathway, while the molecular events prime IRF1 to translocate into nucleus are still obscure. Here we identified STXBP6, previously recognized involving in SNARE complex assembly, negatively regulates PD-L1 transcription via retention of IRF1 in cytoplasm. IFN-γ exposure stimulates accumulation of cytosolic IRF1, which eventually saturates STXBP6 and triggers nuclear translocation of IRF1. Nuclear IRF1 in turn inhibits STXBP6 expression and thereby liberates more IRF1 to migrate to nucleus. Therefore, we identified a novel positive feedback loop between STXBP6 and IRF1 in regulation of PD-L1 expression in cancer. Furthermore, we demonstrate STXBP6 overexpression significantly inhibits T cell activation both in vitro and in vivo. These findings offer new insight into the complexity of PD-L1 expression in cancer and suggest a valuable measure to predict the response to PD-1/PD-L1-based immunotherapy.

Vaxign-ML: supervised machine learning reverse vaccinology model for improved prediction of bacterial protective antigens.

MOTIVATION: Reverse vaccinology (RV) is a milestone in rational vaccine design, and machine learning (ML) has been applied to enhance the accuracy of RV prediction. However, ML-based RV still faces challenges in prediction accuracy and program accessibility. RESULTS: This study presents Vaxign-ML, a supervised ML classification to predict bacterial protective antigens (BPAgs). To identify the best ML method with optimized conditions, five ML methods were tested with biological and physiochemical features extracted from well-defined training data. Nested 5-fold cross-validation and leave-one-pathogen-out validation were used to ensure unbiased performance assessment and the capability to predict vaccine candidates against a new emerging pathogen. The best performing model (eXtreme Gradient Boosting) was compared to three publicly available programs (Vaxign, VaxiJen, and Antigenic), one SVM-based method, and one epitope-based method using a high-quality benchmark dataset. Vaxign-ML showed superior performance in predicting BPAgs. Vaxign-ML is hosted in a publicly accessible web server and a standalone version is also available. AVAILABILITY AND IMPLEMENTATION: Vaxign-ML website at http://www.violinet.org/vaxign/vaxign-ml, Docker standalone Vaxign-ML available at https://hub.docker.com/r/e4ong1031/vaxign-ml and source code is available at https://github.com/VIOLINet/Vaxign-ML-docker. SUPPLEMENTARY INFORMATION: Supplementary data are available at Bioinformatics online.

The combination of PRL-3 inhibitor with sorafenib synergistically promotes AML apoptosis.

Phosphatase of regenerating liver-3 (PRL-3) is recognized as a novel independent crucial driver for AML progression. Thus, the specific inhibitor of PRL-3 would be a potential therapeutic agent to AML in clinics, but there are not enough preclinical applications reported yet. Here we evaluated the cytotoxicity of PRL-3 inhibitor, BR-1, against AML cells ML-1 and MOLM-13. Meanwhile, the effect of BR-1 on the biological characteristics of AML cells and the underlying mechanism was investigated along with the combination of BR-1 and sorafenib on the AML cell viability. Our results show that BR-1 promotes apoptosis by inactivation of the JAK/STAT5 and PI3K/AKT pathways, while inhibits cell proliferation through arresting cell cycle in the S phase. In addition, a combination of BR-1 with sorafenib can further improve the therapeutic effect on AML. Thus, our results demonstrated that BR-1 would be a novel and potent therapeutic agent to AML, and its combination with other anti-AML drugs would be a promising strategy for AML therapy.

Antibiotic fidaxomicin is an RdRp inhibitor as a potential new therapeutic agent against Zika virus.

BACKGROUND: Zika virus (ZIKV) infection is a global health problem, and its complications, including congenital Zika syndrome and Guillain-Barré syndrome, constitute a continued threat to humans. Unfortunately, effective therapeutics against ZIKV infection are not available thus far. METHODS: We screened the compounds collection consisting of 1789 FDA-approved drugs by a computational docking method to obtain anti-ZIKV candidate compounds targeting ZIKV RNA-dependent RNA polymerase (RdRp). SPR (BIAcore) assay was employed to demonstrate the candidate compounds' direct binding to ZIKV RdRp, and polymerase activity assay was used to determine the inhibitory effect on ZIKV RdRp-catalyzed RNA synthesis. The antiviral effects on ZIKV in vitro and in vivo were detected in infected cultured cells and in Ifnar1-/- mice infected by ZIKV virus using plaque assay, western blotting, tissue immunofluorescence, and immunohistochemistry. RESULTS: Here, we report that a first-in-class macrocyclic antibiotic, which has been clinically used to treat Clostridium difficile infection, fidaxomicin, potently inhibits ZIKV replication in vitro and in vivo. Our data showed that fidaxomicin was effective against African and Asian lineage ZIKV in a wide variety of cell lines of various tissue origins, and prominently suppressed ZIKV infection and significantly improved survival of infected mice. In addition, fidaxomicin treatment reduced the virus load in the brains and testes, and alleviated ZIKV-associated pathological damages, such as paralysis, hunching, and neuronal necrosis in the cerebra. Furthermore, our mechanistic study showed that fidaxomicin directly bound ZIKV NS5 protein and inhibited the RNA synthesis-catalyzing activity of ZIKV RdRp. CONCLUSIONS: Our data suggest that fidaxomicin may represent an effective anti-ZIKV agent. In the light that fidaxomicin is already a clinically used drug, there might be a promising prospect in the development of fidaxomicin to be an antiviral therapeutic.

LncRNA HOXA11-AS promotes hepatocellular carcinoma progression by repressing miR-214-3p.

Accumulating studies supported that lncRNAs played important roles in tumorigenesis. LncRNA HOXA11-AS was a novel lncRNA that has been proved to involved in several tumours. However, the role of HOXA11-AS in the development of hepatocellular carcinoma (HCC) remains to be explained. In our study, we showed that HOXA11-AS expression was up-regulated in the HCC tissues, and the higher expression of HOXA11-AS was associated with the advanced stage in the HCC samples. In addition, we indicated that the expression of HOXA11-AS was up-regulated in HCC cell lines (Hep3B, SMMC-7721, MHCC97-H and BEL-7402) compared with normal liver cell lines (HL-7702). Overexpression of HOXA11-AS promoted HCC proliferation and invasion and induced the epithelial-mesenchymal transition (EMT) and knockdown of HOXA11-AS suppressed the HCC cell proliferation and invasion. However, we showed that miR-214-3p expression was down-regulated in the HCC tissues and cell lines. Ectopic expression of miR-214-3p suppressed HCC cell proliferation and invasion. Furthermore, we indicated that overexpression of HOXA11-AS decreased the miR-214-3p expression and the expression of miR-214-3p was negatively related with the HOXA11-AS expression in HCC samples. Ectopic expression of HOXA11-AS increased HCC proliferation and invasion and induced EMT through inhibiting miR-214-3p expression. These data suggested that HOXA11-AS/miR-214-3p axis was responsible for development of HCC.

Expression patterns of SH3BGR family members in zebrafish development.

SH3 domain-binding glutamic acid-rich (SH3BGR) gene family is composed of SH3BGR, SH3BGRL, SH3BGRL2, and SH3BGRL3 which encodes a cluster of small thioredoxin-like proteins and shares a Src homology 3 (SH3) domain. However, biological functions of SH3BGR family members are largely elusive. Given that zebrafish (Danio rerio) sh3bgrl, sh3bgrl2, sh3bgrl3, and sh3bgr are evolutionally identical to their corresponding human orthologues, we analyzed the spatiotemporal expression of SH3BGR family members in zebrafish embryonic development stages by in situ hybridization. Our results revealed that except sh3bgrl, other members are all maternally expressed, especially for sh3bgrl3 that is strongly expressed from one-cell stage to juvenile fishes. In situ expression patterns of SH3BGR members are similar in the very early developmental stages, including with commonly strong expression in intestines, olfactory bulbs, and neuromasts for neural system building up. Organ-specific expressions are also demonstrated, of which sh3bgr is uniquely expressed in sarcomere, and sh3bgrl3 in liver. sh3bgrl and sh3bgrl2 are similarly expressed in intestines, notochords, and neuromasts after 12-h post-fertilization of embryos. Eventually, messenger RNAs (mRNAs) of all sh3bgr members are mainly constrained into intestines of juvenile fishes. Collectively, our study clarified the expression patterns of sh3bgr family members in diverse organogenesis in embryonic development and indicates that SH3BGR members may play predominant roles in neural system development and in maintenance of normal function of digestive organs, especially for intestine homeostasis. However, their expression patterns are varied with the development stages and organ types, suggesting that the aberrant expression of these members would result in multiple diseases.

PRL-3 promotes migration and invasion and is associated with poor prognosis in salivary adenoid cystic carcinoma.

BACKGROUND: PRL-3 had been found to be involved in tumorigenesis in various malignancies. In this study, we investigated the role of PRL-3 in the development, migration, and invasion of salivary adenoid cystic carcinoma (SACC). METHODS: Immunohistochemistry (IHC) was used to analyze the role of PRL-3 in the development and prognosis of SACC. Then, we overexpressed or inhibited the expression of PRL-3 in paired SACC cells to analyze the role of PRL-3 in the migration and invasion of SACC. In vitro migration and invasion assays were used. Western blotting was used to detect metastasis-related protein levels. RESULTS: IHC results confirmed that the deregulation of PRL-3 was a frequent event in SACC; the upregulation of PRL-3 was related to clinical stages, vital status, and distant metastasis, which was associated with reduced overall survival and disease-free survival. SACC-LM cells with higher migratory and invasive abilities had more robust PRL-3 protein expression than SACC-83 cells with lower migratory and invasive abilities. PRL-3 overexpression promoted cell migration, invasion, and proliferation, led to simultaneous upregulation of phosphorylated PRL-3, pERK1/2, Slug, vimentin, and downregulation of E-cadherin in SACC-83 cells. However, the inhibition of PRL-3 by PRL-3 inhibitor or PRL-3 siRNA in SACC-LM cells inhibited cell migration, invasion, and proliferation, resulted in simultaneous downregulation of phosphorylated PRL-3, pERK1/2, Slug, vimentin, and upregulation of E-cadherin. CONCLUSIONS: Our results confirm that PRL-3 plays an important role in the development of SACC and contributes to the migratory and invasive abilities of SACC.

STAT1 inhibits human hepatocellular carcinoma cell growth through induction of p53 and Fbxw7.

BACKGROUND: Aberrant STAT1 signaling is observed in human hepatocellular carcinoma (HCC) and has been associated with the modulation of cell proliferation and survival. However, the role of STAT1 signaling in HCC and its underlying mechanism remain elusive. METHODS: We transiently transfected pcDNA3.1-STAT1 and STAT1 siRNA into SMMC7721 and HepG2 cells. Western blot and qRT-PCR examined the expression of protein and RNA of target genes. Cell viability was assessed using MTT assay, and cell cycle and apoptosis were analyzed by flow cytometry. RESULTS: We found that STAT1 overexpression increased protein expression of p53 and Fbxw7, and downregulated the expression of cyclin A, cyclin D1, cyclin E, CDK2, Hes-1 and NF-κB p65. These changes led to growth inhibition and induced G0/G1 cell cycle arrest and apoptosis in SMMC7721 and HepG2 cells. Conversely, ablation of STAT1 had the opposite effect on p53, Fbxw7, Hes-1, NF-κB p65, cyclin A, cyclin D1, cyclin E and CDK2, and improved the viability of SMMC7721 and HepG2 cells. CONCLUSIONS: Our data indicate that STAT1 exerts tumor-suppressive effects in hepatocarcinogenesis through induction of G0/G1 cell cycle arrest and apoptosis, and may provide a basis for the design of new therapies for the intervention of HCC in the clinic.

Independent oncogenic and therapeutic significance of phosphatase PRL-3 in FLT3-ITD-negative acute myeloid leukemia.

BACKGROUND: Internal tandem duplication of FMS-like tyrosine kinase (FLT3-ITD) is well known to be involved in acute myeloid leukemia (AML) progression, but FLT3-ITD-negative AML cases account for 70% to 80% of AML, and the mechanisms underlying their pathology remain unclear. This study identifies protein tyrosine phophatase PRL-3 as a key mediator of FLT3-ITD-negative AML. METHODS: A total of 112 FLT3-ITD-negative AML patients were sampled between 2010 and 2013, and the occurrence of PRL-3 hyperexpression in FLT3-ITD-negative AML was evaluated by multivariate probit regression analysis. Overexpression or depletion of endogenous PRL-3 expression with the specific small interfering RNAs was performed to investigate the role of PRL-3 in AML progression. Xenograft models were also used to confirm the oncogenic role of PRL-3. RESULTS: Compared to healthy donors, PRL-3 is upregulated more than 3-fold in 40.2% of FLT3-ITD-negative AML patients. PRL-3 expression level is adversely correlated to the overall survival of the AML patients, and the AML relapses accompany with re-upregulation of PRL-3. Mechanistically, aberrant PRL-3 expression promoted cell cycle progression and enhanced the antiapoptotic machinery of AML cells to drug cytotoxicity through downregulation of p21 and upregulation of Cyclin D1 and CDK2 and activation of STAT5 and AKT. Depletion of endogenous PRL-3 sensitizes AML cells to therapeutic drugs, concomitant with apoptosis by upregulation of cleaved PARP (poly ADP ribose polymerase) and apoptosis-related caspases. Xenograft assays further confirmed PRL-3's oncogenic role in leukemogenesis. CONCLUSIONS: Our results demonstrated that PRL-3 is a novel independent crucial player in both FLT3-ITD-positive and FLT3-ITD-negative AML and could be a potential therapeutic target.

RBP-J-interacting and tubulin-associated protein induces apoptosis and cell cycle arrest in human hepatocellular carcinoma by activating the p53-Fbxw7 pathway.

Aberrant Notch signaling is observed in human hepatocellular carcinoma (HCC) and has been associated with the modulation of cell growth. However, the role of Notch signaling in HCC and its underlying mechanism remain elusive. RBP-J-interacting and tubulin-associated (RITA) mediates the nuclear export of RBP-J to tubulin fibers and downregulates Notch-mediated transcription. In this study, we found that RITA overexpression increased protein expression of p53 and Fbxw7 and downregulated the expression of cyclin D1, cyclin E, CDK2, Hes-1 and NF-κB p65. These changes led to growth inhibition and induced G0/G1 cell cycle arrest and apoptosis in SMMC7721 and HepG2 cells. Our findings indicate that RITA exerts tumor-suppressive effects in hepatocarcinogenesis through induction of G0/G1 cell cycle arrest and apoptosis and suggest a therapeutic application of RITA in HCC.

Inhibitory effect of baicalin on collagen-induced arthritis in rats through the nuclear factor-κB pathway.

This study focused on the potential therapeutic effect of baicalin on collagen-induced arthritis (CIA) in rats and the underlying mechanisms. The CIA rats were injected with baicalin (50, 100, or 200 mg/kg) once daily for 30 days. The rats were monitored for clinical severity of arthritis, and joint tissues were used for radiographic assessment and histologic examination. We quantified tumor necrosis factor-α (TNF-α) and interleukin-1ß (IL-1ß) in experimental animals and used Western blots to assess levels of protein abundance, phosphorylation, and acetylation of nuclear factor (NF)-κB p65 and sirtuin 1 (sirt1) protein expression in joint tissues. Human fibroblast-like synoviocytes from rheumatoid arthritis (HFLS-RA) were adopted in further mechanistic investigations. Baicalin intraperitoneal injection for 30 days dose-dependently blocked clinical manifestations of CIA, such as functional impairment and swollen red paws. Meanwhile, it alleviated collagen-induced joint inflammation injury and inhibited the secretion of TNF-α and IL-1ß in both rat synovium and HFLS-RA. Further mechanistic investigations revealed that baicalin suppresses NF-κB p65 protein expression and phosphorylation in synovial tissue and human-derived synoviocytes. Moreover, the acetylation of NF-κB p65 was downregulated by baicalin, which negatively correlates with the baicalin-induced upregulation of sirt1 expression in the same conditions. The data indicate that CIA in rats can be alleviated by baicalin treatment via relieving joint inflammation, which is related to the suppression of synovial NF-κB p65 protein expression and the elevation of its deacetylation by sirt1.

PRL-3 suppresses c-Fos and integrin α2 expression in ovarian cancer cells.

BACKGROUND: Phosphatase of regenerating liver-3 (PRL-3), a protein tyrosine phosphatase, is highly expressed in multiple human cancers and strongly implicated in tumor progression and cancer metastasis. However, the mechanisms by which PRL-3 promotes cancer cell migration, invasion, and metastasis are not very well understood. In this study, we investigated the contribution and molecular mechanisms of PRL-3 in ovarian cancer progression. METHODS: PRL-3 protein expression was detected on ovarian cancer tissue microarrays using immunohistochemistry. Stable PRL-3 depleted cell lines were generated using short hairpin RNA (shRNA) constructs. The migration and invasion potential of these cells were analyzed using Transwell and Matrigel assays, respectively. Immunoblotting and immunofluorescence were used to detect protein levels and distribution in PRL-3-ablated cells and the control cells. Cell morphology was observed with hematoxylin-eosin staining and transmission electron microscopy. Finally, PRL-3-ablated and control cells were injected into nude mice for xenograft tumorigenicity assays. RESULTS: Elevated PRL-3 expression was detected in 19% (26 out of 135) of human ovarian cancer patient samples, but not in normal ovary tissues (0 out of 14). Stable depletion of PRL-3 in A2780 ovarian cancer cells resulted in decreased migration ability and invasion activity compared with control parental A2780 cells. In addition, PRL-3-ablated cells also exhibited flattened morphology and extended lamellipodia. To address the possible molecular basis for the altered phenotypes associated with PRL-3 down-regulation, we assessed the expression profiles of various proteins involved in cell-matrix adhesion. Depletion of PRL-3 dramatically enhanced both RNA and protein levels of the cell surface receptor integrin α2, but not its heterologous binding partner integrin ß1. Inhibition of PRL-3 also correlated with elevated expression and phosphorylation of paxillin. A pronounced increase in the expression and activation of c-fos, a transcriptional activator of integrin α2, was observed in these PRL-3 knock-down cells. Moreover, forced expression of EGFP-PRL-3 resulted in the suppression of both integrin α2 and c-fos expression in A2780 cells. Significantly, using a xenograft tumor model, we observed a greatly reduced tumorigenicity of A2780 PRL-3 knock-down cells in vivo. CONCLUSIONS: These results suggest that PRL-3 plays a critical role in ovarian cancer tumorigenicity and maintaining the malignant phenotype. PRL-3 may inhibit c-fos transcriptional regulation of integrin α2 signaling. Our results strongly support a role for PRL-3 as a promising therapeutic target and potential early biomarker in ovarian cancer progression.

RITA inhibits growth of human hepatocellular carcinoma through induction of apoptosis.

RBP-J-interacting and tubulin-associated (RITA) is a novel RBP-J-interacting protein that downregulates Notch-mediated transcription. The current study focuses on the antitumor effect of RITA in human hepatocellular carcinoma (HCC) and aims to explore its molecular mechanism. Thirty paired HCC and adjacent non-tumoral liver samples were analyzed by real-time quantitative reverse transcriptase polymerase chain reaction (qRT-PCR). RITA overexpression was induced by transfection of a pcDNA3.1-Flag-RITA plasmid into HepG2 cells. RITA knockdown was achieved by siRNA transfection. mRNA and protein expression of target genes were quantified by qRT-PCR and Western blotting, respectively. Cell proliferation and apoptosis were measured using MTT assay and flow cytometry. Our results demonstrate that adjacent nontumoral liver samples exhibited increased RITA expression compared to HCC tissues (p < 0.05); RITA levels were associated with tumor differentiation status. Overexpression of RITA suppressed cell proliferation and promoted early apoptosis, while its silencing promoted cell growth dramatically (p < 0.05). RITA overexpression upregulated p53 and reduced cyclin E levels, whereas silencing of RITA had the opposite effect on p53 and cyclin E expression. Our in vitro results represent the first evidence that RITA might suppress tumor growth and induce apoptosis in HCCs, and may be a potent antitumoral agent for HCC treatment that deserves further exploration.

Integrative analysis of multi-omics data reveals inhibition of RB1 signaling promotes apatinib resistance of hepatocellular carcinoma.

Although apatinib is a promising drug for the treatment of liver cancer, the underlying drug resistance mechanism is still unclear. Here, we constructed apatinib-resistant HepG2 cells. We then characterized the epigenomic, transcriptomic, and proteomic landscapes both in apatinib-resistant and non-resistant HepG2 cells. Differential expression, ATAC-seq, and proteomic data analyses were performed. We found that the cell cycle related protein RB1 may play an essential role in the process of apatinib resistant to hepatocarcinoma. Moreover, there were extensive variations at the transcriptome, epigenetic, and proteomic level. Finally, quantitative PCR (qPCR) and western blot analysis showed that expression level of RB1 in apatinib-resistant cell as well as the samples of patients in progressive disease were significantly lower than that in controls. Those results also showed that the RB1 pathway inhibitors CDK2-IN-73 and Palbociclib could relieve the resistance of apatinib resistant cells. Our results further enhance our understanding of the anti-tumorigenic and anti-angiogenic efficacy of apatinib in liver cancer and provide a novel perspective regarding apatinib resistance. Furthermore, we proved that CDKN2B inhibition of RB1 signaling promoted apatinib resistance in hepatocellular carcinoma. Those findings have greatly important biological significance for the resistance of apatinib and the treatment of liver cancer.

Loss of the adaptor protein Sh3bgrl initiates ovarian fibrosis in zebrafish.

Ovarian fibrosis is a reproduction obstacle leading to female infertility in vertebrates, but the cause underlying the cellular events is unclear. Here, we found that the small adaptor protein SH3-domain-binding glutamate-rich protein like (Sh3bgrl) plays an important role in female reproduction in zebrafish. Two sh3bgrl mutant alleles that result in sh3bgrl depletion contribute to female spawning inability. Comparative transcriptome analysis revealed that sh3bgrl knockout mechanistically causes the upregulation of genes associated with extracellular matrix (ECM) and fiber generation in the zebrafish ovary. Consequently, extra ECM or fibers accumulate and are deposited in the ovary, resulting in eventual spawning inability. Our findings thus provide insights into understanding the underlying mechanism of infertility by ovarian fibrosis and provide a novel and valuable model to study female reproduction abnormality.

SH3BGRL Suppresses Liver Tumor Progression through Enhanced ATG5-Dependent Autophagy.

SH3BGRL, an adaptor protein, is upregulated in breast cancers and indicates its tumorigenic role. But the function of SH3BGRL in other types of cancers is largely unknown. Here, we modulate SH3BGRL expression level in two liver cancer cells and conduct both in vitro and in vivo analyses of SH3BGRL in cell proliferation and tumorigenesis. Results demonstrate that SH3BGRL notably inhibits cell proliferation and arrests the cell cycle in both LO2 and HepG2 cells. Molecularly, SH3BGRL upregulates the expression of ATG5 from proteasome degradation as well as the inhibitions of Src activation and its downstream ERK and AKT signaling pathways, which eventually enhance autophagic cell death. The xenograft mouse model reveals that SH3BGRL overexpression can efficiently suppress tumorigenesis in vivo, while the additional silencing ATG5 in SH3BGRL-overexpressing cells attenuates the inhibitory effect of SH3BGRL on both hepatic tumor cell proliferation and tumorigenicity in vivo. The relevance of SH3BGRL downregulation in liver cancers and their progression is validated based on the large-scale tumor data. Taken together, our results clarify the suppressive role of SH3BGRL in tumorigenesis of liver cancer, which would be of help to the diagnosis of liver cancer, while either promoting the autophagy of liver cancer cells or inhibiting the downstream signaling induced from SH3BGRL downregulation would be a promising therapy.