Serine and arginine rich splicing factor 3 (SRSF3), an SR-rich family protein, has an oncogenic function in various kinds of cancer. However, the detailed mechanism of the function had not been previously clarified. Here, we showed that the SRSF3 splicer regulated the expression profile of the pyruvate kinase, which is one of the rate-limiting enzymes in glycolysis. Most cancer cells express pyruvate kinase muscle 2 (PKM2) dominantly to maintain a glycolysis-dominant energy metabolism. Overexpression of SRSF3, as well as that of another splicer, polypyrimidine tract binding protein 1 (PTBP1) and heterogeneous nuclear ribonucleoprotein A1 (hnRNPA1), in clinical cancer samples supported the notion that these proteins decreased the Pyruvate kinase muscle 1 (PKM1)/PKM2 ratio, which positively contributed to a glycolysis-dominant metabolism. The silencing of SRSF3 in human colon cancer cells induced a marked growth inhibition in both in vitro and in vivo experiments and caused an increase in the PKM1/PKM2 ratio, thus resulting in a metabolic shift from glycolysis to oxidative phosphorylation. At the same time, the silenced cells were induced to undergo autophagy. SRSF3 contributed to PKM mRNA splicing by co-operating with PTBP1 and hnRNPA1, which was validated by the results of RNP immunoprecipitation (RIP) and immunoprecipitation (IP) experiments. These findings altogether indicated that SRSF3 as a PKM splicer played a positive role in cancer-specific energy metabolism.

MicroRNAs (miRNAs) are small RNAs present in extracellular vesicles (EVs) that, when transferred to a target cell, affect its biological functions. Plant miRNAs regulate the expression of certain mammalian genes. Here, we characterized EVs in fruit and vegetable juice, and their miRNA cargo, and investigated whether such miRNA-containing EVs could be taken up by mammalian enterocytes in vitro. Using filtration and ultra-centrifugation methods, EVs were purified from commercially available and manually squeezed plant juice. EV morphological features and subcellular localization were analyzed using the NanoSight tracking system and electron microscopy. Plant EV miRNA levels were evaluated using quantitative reverse transcription PCR. For the in vitro EV uptake experiments, rat intestinal epithelial cells (IEC6) were used. Plant EVs shared morphological features with mammalian EVs and contained miR156a-5p, miR166a-3p, and miR168a-5p. EVs were present in the cell sap-filled central vacuoles and were taken up by IEC6 cells. Edible plant cells produce EVs that contain various miRNAs and release them into the central vacuole. The exogenous plant EVs are taken up by mammalian enterocytes in vitro. These findings suggest the possibility that exogenous plant miRNAs carried by EVs can be absorbed via the gastrointestinal tract.

Macro-autophagy (autophagy) is a highly conserved eukaryotic intracellular process of self-digestion caused by lysosomes on demand, which is upregulated as a survival strategy upon exposure to various stressors, such as metabolic insults, cytotoxic drugs, and alcohol abuse. Paradoxically, autophagy dysfunction also contributes to cancer and aging. It is well known that regulating autophagy by targeting specific regulatory molecules in its machinery can modulate multiple disease processes. Therefore, autophagy represents a significant pharmacological target for drug development and therapeutic interventions in various diseases, including cancers. According to the framework of autophagy, the suppression or induction of autophagy can exert therapeutic properties through the promotion of cell death or cell survival, which are the two main events targeted by cancer therapies. Remarkably, natural products have attracted attention in the anticancer drug discovery field, because they are biologically friendly and have potential therapeutic effects. In this review, we summarize the up-to-date knowledge regarding natural products that can modulate autophagy in various cancers. These findings will provide a new position to exploit more natural compounds as potential novel anticancer drugs and will lead to a better understanding of molecular pathways by targeting the various autophagy stages of upcoming cancer therapeutics.

Bladder cancer is one of the most difficult malignancies to control. We explored the use of a novel RNA-interference method for a driver oncogene regulating cancer specific energy metabolism by the combination treatment with a small interfering RNA (siRNA) and a microRNA. After transfection of T24 and 253JB-V cells with miR-145 and/or siR-PTBP1, we examined the effects of cell growth and gene expression by performing the trypan blue dye exclusion test, Western blot, Hoechst 33342 staining, reverse transcription polymerase chain reaction (RT-PCR), and electron microscopy. The anti-cancer effects of xenograft model mice with miR-145 and/or siR-PTBP1 were then assessed. The combination treatment induced the deeper and longer growth inhibition and reduced the levels of both mRNA and protein expression of c-Myc and polypyrimidine tract-binding protein 1 (PTBP1) more than each single treatment. Notably, the combination treatment not only impaired the cancer specific energy metabolism by inhibiting c-Myc/PTBP1/PKMs axis but also inactivated MAPK/ERK and PI3K/AKT pathways examined in vitro and in vivo. Furthermore, the combination treatment induced apoptosis or autophagy; but, in some cells, apoptotic cell death was accompanied by autophagy, because the condensation of chromatin and many autophagosomes were coexistent. This combination treatment could be a novel RNA-interference strategy through the systemic silencing of the Warburg effect-promoting driver oncogene PTBP1 in bladder cancer cells.

Mitochondrial electron transport chain complex I (ETCC1) is the essential core of cancer metabolism, yet potent ETCC1 inhibitors capable of safely suppressing tumor growth and metastasis in vivo are limited. From a plant extract screening, we identified petasin (PT) as a highly potent ETCC1 inhibitor with a chemical structure distinct from conventional inhibitors. PT had at least 1700 times higher activity than that of metformin or phenformin and induced cytotoxicity against a broad spectrum of tumor types. PT administration also induced prominent growth inhibition in multiple syngeneic and xenograft mouse models in vivo. Despite its higher potency, it showed no apparent toxicity toward nontumor cells and normal organs. Also, treatment with PT attenuated cellular motility and focal adhesion in vitro as well as lung metastasis in vivo. Metabolome and proteome analyses revealed that PT severely depleted the level of aspartate, disrupted tumor-associated metabolism of nucleotide synthesis and glycosylation, and downregulated major oncoproteins associated with proliferation and metastasis. These findings indicate the promising potential of PT as a potent ETCC1 inhibitor to target the metabolic vulnerability of tumor cells.

This study was conducted to investigate the autophagic response of Sertoli cells (SCs) to acute ethanol toxicity using in vivo and in vitro models. Adult Wistar rats were intraperitoneally injected with either 5 g/kg ethanol or phosphate-buffered saline (for the control group) and sacrificed 0, 3, 6 and 24 h after injection. Compared to the control group, enhanced germ cell apoptosis was observed in the ethanol-treated rats (ETRs) in association with upregulation of iNOS and reduced expression of androgen receptor protein levels in SCs, which were resistant to apoptosis. Meanwhile, autophagy was upregulated in ETR SCs (peaking at 24 h) compared to the control group, as evidenced by transcription factor EB (TFEB) nuclear translocation, enhanced expression of microtubule-associated protein 1 light chain3-II (LC3-II), lysosome-associated membrane protein-2 (LAMP-2), pan cathepsin protein levels and reduced expression of p62. This upregulation of SC autophagy was confirmed ultrastructurally by enhanced formation of autophagic vacuoles and by immunofluorescent double labelling of autophagosomal and lysosomal markers. Study of cultured SCs confirmed enhanced autophagic response to ethanol toxicity, which was cytoprotective based on decreased viability of SCs upon blocking autophagy with 3-methyladenine (3-MA). The results highlighted the molecular mechanisms of prosurvival autophagy in ETR SCs for the first time, and may have significant implications for male fertility.