No abstract

Autophagy plays diverse roles in Ras-related tumorigenesis. H-ras(val12) induces autophagy through multiple signaling pathways including Raf-1/ERK pathway, and various ERK downstream molecules of autophagy have been reported. In this study, Bcl-2/adenovirus E1B 19-kDa-interacting protein 3 (BNIP3) is identified as a downstream transducer of the Ras/Raf/ERK signaling pathway to induce autophagy. BNIP3 was upregulated by H-ras(val12) at the transcriptional level to compete with Beclin 1 for binding with Bcl-2. H-ras(val12)-induced autophagy suppresses cell proliferation demonstrated both in vitro and in vivo by expression of ectopic BNIP3, Atg5, or interference RNA of BNIP3 (siBNIP3) and Atg5 (shAtg5) using mouse NIH3T3 and embryo fibroblast cells. H-ras(val12) induces different autophagic responses depending on the duration of Ras overexpression. After a short time (48 hours) of Ras overexpression, autophagy inhibits cell proliferation. In contrast, a longer time (2 weeks) of Ras overexpression, cell proliferation was enhanced by autophagy. Furthermore, overexpression of mutant Ras, BNIP3, and LC3-II was detected in bladder cancer T24 cells and the tumor parts of 75% of bladder cancer specimens indicating a positive correlation between autophagy and tumorigenesis. Taken together, our mouse model demonstrates a balance between BNIP3-mediated autophagy and H-ras(val12)-induced tumor formation and reveals that H-ras(val12) induces autophagy in a BNIP3-dependent manner, and the threshold of autophagy plays a decisive role in H-ras(val12)-induced tumorigenesis. Our findings combined with others' reports suggest a new therapeutic strategy against Ras-related tumorigenesis by negative or positive regulation of autophagic activity, which is determined by the level of autophagy and tumor progression stages.

Ferroptosis, a recently discovered form of iron-dependent cell death, requires an increased level of lipid-reactive oxygen species (ROS). Ferritinophagy, a ferritin degradation pathway, depends on a selective autophagic cargo receptor (NCOA4). By screening various types of natural compounds, formosanin C (FC) was identified as a novel ferroptosis inducer, characterized by attenuations of FC-induced viability inhibition and lipid ROS formation in the presence of ferroptosis inhibitor. FC also induced autophagic flux, evidenced by preventing autophagic marker LC3-II degradation and increasing yellow LC3 puncta in tandem fluorescent-tagged LC3 (mRFP-GFP) reporter plasmid (ptfLC3) transfected cells when combined with autophagic flux inhibitor. It is noteworthy that FC-induced ferroptosis and autophagic flux were stronger in HepG2 cells expressing higher NCOA4 and lower ferritin heavy chain 1 (FTH1) levels, agreeing with the results of gene expression analysis using CTRP and PRISM, indicating that FTH1 expression level exhibited a significant negative correlation with the sensitivity of the cells to a ferroptosis inducer. Confocal and electron microscopy confirmed the pronounced involvement of ferritinophagy in FC-induced ferroptosis in the cells with elevated NCOA4. Since ferroptosis is a non-apoptotic form of cell death, our data suggest FC has chemotherapeutic potential against apoptosis-resistant HCC with a higher NCOA4 expression via ferritinophagy.

Approximately 350 million people worldwide are chronically infected by hepatitis B virus (HBV). HBV causes severe liver diseases including cirrhosis and hepatocellular carcinoma (HCC). In about 25% of affected patients, HBV infection proceeds to HCC. Therefore, the mechanisms by which HBV affects the host cell to promote viral replication and its pathogenesis have been the subject of intensive research efforts. Emerging evidence indicates that both autophagy and microRNAs (miRNAs) are involved in HBV replication and HBV-related hepatocarcinogenesis. In this review, we summarize how HBV induces autophagy, the role of autophagy in HBV infection, and HBV-related tumorigenesis. We further discuss the emerging roles of miRNAs in HBV infection and how HBV affects miRNAs biogenesis. The accumulating knowledge pertaining to autophagy and miRNAs in HBV replication and its pathogenesis may lead to the development of novel strategies against HBV infection and HBV-related HCC tumorigenesis.

Curcumin from the rhizome of the Curcuma longa plant has been noted for its chemo-preventative and chemo-therapy activities, and it inhibits the growth of many types of human cancer cell lines. In this study, the mechanisms of cell death involved in curcumin-induced growth inhibition, including cell cycle arrest and induction of apoptosis in human tongue cancer SCC-4 cells, were investigated. Herein, we observed that curcumin inhibited cell growth of SCC-4 cells and induced cell death in a dose-dependent manner. Treatment of SCC-4 cells with curcumin caused a moderate and promoted the G(2) /M phase arrest, which was accompanied with decreases in cyclin B/CDK1 and CDC25C protein levels. Moreover, curcumin significantly induced apoptosis of SCC-4 cells with a decrease of the Bcl-2 level, reduction of mitochondrial membrane potential (ΔΨ(m) ), and promoted the active forms of caspase-3. Curcumin also promoted the releases of AIF and Endo G from the mitochondria in SCC-4 cells by using confocal laser microscope. Therefore, we suggest that curcumin induced apoptosis through a mitochondria-dependent pathway in SCC-4 cells. In addition, we also found that curcumin-induced apoptosis of SCC-4 cells was partly through endoplasmic reticulum stress. In conclusion, curcumin increased G(2) /M phase arrest and induced apoptosis through ER stress and mitochondria-dependent pathways in SCC-4 cells.

Our previous reports showed that justicidin A (JA), a novel and pure arylnaphthalide lignan isolated from Justicia procumbens, induces apoptosis of human colorectal cancer cells and hepatocellular carcinoma cells, leading to the suppression of both tumor cell growth in NOD-SCID mice. Here, we reveal that JA induces autophagy in human colorectal cancer HT-29 cells by conversion of autophagic marker LC3-I to LC3-II. Furthermore, LC3 puncta and autophagic vesicle formation, and SQSTM1/p62 suppression were observed. Administration of autophagy inhibitor (bafilomycin A1 and chloroquine) and transfection of a tandem fluorescent-tagged LC3 (mRFP-GFP) reporter plasmid (ptfLC3) demonstrated that JA induces autophagy flux in HT-29 cells. Expression of LC3, SQSTM1, Beclin 1, and nuclear DNA double-strand breaks (representing apoptosis) were also detected in the tumor tissue of HT-29 cells transplanted into NOD-SCID mice orally administrated with JA. In addition, the expression of autophagy signaling pathway-related molecules p-PDK1, p-mTOR, p-p70S6k/p-RPS6KB2 was decreased, whereas that of class III PI3K, Beclin 1, Atg5-Atg12, and mitochondrial BNIP3 was increased in response to JA. Pre-treatment of the cells with class III PI3K inhibitor 3-methyladenine or Atg5 shRNA attenuated JA-induced LC3-II expression and LC3 puncta formation, indicating the involvement of class III PI3K and Atg5. A novel mechanism was demonstrated in the anticancer compound JA; pre-treatment with 3-methyladenine or Atg5 shRNA blocked JA-induced suppression in cell growth and colony formation, respectively, via inhibition of apoptosis. In contrast, administration of apoptosis inhibitor Z-VAD did not affect JA-induced autophagy. Our data suggest the chemotherapeutic potential of JA for treatment of human colorectal cancer.

No abstract

We previously reported that dengue virus (DENV)‐induced autophagy plays a promoting role in viral replication and pathogenesis both in vitro and in vivo. Although it is known that DENV infection increases glycolysis, which promotes viral replication, the role of glucose metabolism together with autophagic activity in DENV replication remains unclear. In this study, we reveal that DENV2 infection increased autophagic activity, glucose uptake, protein levels of glucose transporter‐1 (GLUT1), and glycolysis rate‐limiting enzyme hexokinase‐2 (HK2) in cells. Furthermore, the protein levels of LC3‐II and HK2 were increased in the brain tissues of the DENV2‐infected suckling mice. However, DENV2 infection decreased ATP level and showed no effect on mRNA expression of HK2 and phosphofructokinase, as well as lactate production, indicating that DENV2‐regulated glycolytic flux occurs at the post‐transcriptional level and is lactate pathway‐independent. Moreover, amiodarone‐induced autophagic activity, glucose uptake, HK2 level, and viral titer were reversed by the autophagy inhibitor spautin‐1 or silencing of Atg5 gene expression. Intriguingly, blocking of glycolysis, HK2 protein level, and viral titer were accordingly decreased, but autophagic activity was increased, suggesting the existence of another regulation mechanism that influences the relationship between glycolysis and autophagy. This is the first report to reveal that DENV2‐induced autophagy positively regulates glycolysis and viral replication in vitro and in vivo. Our findings open a new avenue wherein metabolic modulation could be used as a target for the treatment of DENV infection.