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In recent years, significant attention has focused on circular RNA (circRNA) translation to determine its clinical significance. Cap-independent translation of circRNAs driven by an internal ribosome entry site (IRES) or an N6-methyladenosine (m

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PDF file - 5906K, Supplemental Table I - Inhibition of mammary carcinogenesis in Wistar-Furth rats by mda-7/IL-24 Supplemental Table II - Microarray analysis of differential gene expression in FE1.2 + IL-24S compared to FE1.2 + IL-24AS cells. Supplemental Figure 1: Serum starvation induces GAS3 expression in both FE1.2 and FE1.3 cells. Supplemental Figure 2: GAS3 downregulation decreases the growth of cells in soft agar. Supplemental Figure 3: STAT3 inhibition reduced attachment of attaches cells to culture plates. Supplemental Figure 4: GAS3 expression reduced attachment of confluent cells to culture plates. Supplemental Figure 5: Binding of GAS3 to β1 integrin using immunoprecipitation assay. Supplemental Figure 6: GAS3 expression inhibits attachment to and proliferation on fibronectin coated culture plates. Supplemental Figure 7: GAS3 expression increases attachment to laminin coated culture plates. Supplemental Figure 8: GAS3 expression by immunohistochemistry.

Melanoma differentiation-associated gene (MDA)-7)/interleukin (IL)-24, a member of the IL-10 family of cytokines, inhibits growth of various human cancer cells, yet the underlying mechanism is largely unknown. Here, we report that mda-7/IL-24 efficiently suppresses the development of rat mammary tumors in vivo. Microarray analysis for genes differentially expressed in rat mammary tumor cells overexpressing MDA-7/IL-24 compared with those that do not express this cytokine identified growth arrest-specific gene-3 (gas3) as a target for mda-7/IL-24. Upregulation of gas3 by mda-7/IL-24 was STAT3 dependent. Induction of gas3 inhibited attachment and proliferation of tumor cells in vitro and in vivo by inhibiting the interaction of β1 integrin with fibronectin. A mutated GAS3, which is unable to bind β1 integrin, was also unable to inhibit fibronectin-mediated attachment and cell growth both in adherent and suspension cultures, suggesting that GAS3 exerts its effects through interaction with and regulation of β1 integrin. Thus, mda-7/IL-24 inhibits breast cancer growth, at least in part, through upregulation of GAS3 and disruption of β1 integrin function. Importantly, the expression of the mda-7/IL-24 receptor, IL-20R1, is highly correlated with GAS3 expression in human breast cancer (P = 1.02 × 10−9), and the incidence of metastases is significantly reduced in patients with HER2+ breast cancer expressing high-levels of IL-20R1. Together, our results identify a novel MDA-7/IL-24-GAS3-β1integrin–fibronectin signaling pathway that suppresses breast cancer growth and can be targeted for therapy. Mol Cancer Res; 11(6); 593–603. ©2013 AACR.

A novel TRIM family member, TRIM59 gene was characterized to be upregulated in SV40 Tag oncogene–directed transgenic and knockout mouse prostate cancer models as a signaling pathway effector. We identified two phosphorylated forms of TRIM59 (p53 and p55) and characterized them using purified TRIM59 proteins from mouse prostate cancer models at different stages with wild-type mice and NIH3T3 cells as controls. p53/p55-TRIM59 proteins possibly represent Ser/Thr and Tyr phosphorylation modifications, respectively. Quantitative measurements by ELISA showed that the p-Ser/Thr TRIM59 correlated with tumorigenesis, whereas the p-Tyr-TRIM59 protein correlated with advanced cancer of the prostate (CaP). The function of TRIM59 was elucidated using short hairpin RNA (shRNA)-mediated knockdown of the gene in human CaP cells, which caused S-phase cell-cycle arrest and cell growth retardation. A hit-and-run effect of TRIM59 shRNA knockdown was observed 24 hours posttransfection. Differential cDNA microarrray analysis was conducted, which showed that the initial and rapid knockdown occurred early in the Ras signaling pathway. To confirm the proto-oncogenic function of TRIM59 in the Ras signaling pathway, we generated a transgenic mouse model using a prostate tissue–specific gene (PSP94) to direct the upregulation of the TRIM59 gene. Restricted TRIM59 gene upregulation in the prostate revealed the full potential for inducing tumorigenesis, similar to the expression of SV40 Tag, and coincided with the upregulation of genes specific to the Ras signaling pathway and bridging genes for SV40 Tag–mediated oncogenesis. The finding of a possible novel oncogene in animal models will implicate a novel strategy for diagnosis, prognosis, and therapy for cancer. Mol Cancer Ther; 10(7); 1229–40. ©2011 AACR.

PDF file - 269K, Figure S1. The mammosphere cells expressed high levels of versican V1, Sca-1, Sox2, and ALDH1. Figure S2. Versican G3 enhanced breast cancer colony formation. Figure S3. Expression of versican G3 domain promoted breast cancer cells expression of cell self-renewal markers.

Supplementary Methods, Figures 1-4, Tables 1-2 from Characterization of the Oncogenic Activity of the Novel TRIM59 Gene in Mouse Cancer Models

Grouping of tumour samples in TCGA ovarian cancer database for the analysis of FOXA2 and VCAN expression

Table S1 to S3. Description of clinical samples; sequences of primers, siRNAs, and microRNA mimics and inhibitors; and analysis of top regulated genes by mir-590.

miRNAs play important roles in gene regulation, and their dysregulation is associated with many diseases, including epithelial ovarian cancer (EOC). In this study, we determined the expression and function of miR-590-3p in EOC. miR-590-3p levels were higher in high-grade carcinoma when compared with low-grade or tumors with low malignant potential. Interestingly, plasma levels of miR-590-3p were significantly higher in patients with EOC than in subjects with benign gynecologic disorders. Transient transfection of miR-590-3p mimics or stable transfection of mir-590 increased cell proliferation, migration, and invasion. In vivo studies revealed that mir-590 accelerated tumor growth and metastasis. Using a cDNA microarray, we identified forkhead box A2 (FOXA2) and versican (VCAN) as top downregulated and upregulated genes by mir-590, respectively. miR-590-3p targeted FOXA2 3′ UTR to suppress its expression. In addition, knockdown or knockout of FOXA2 enhanced cell proliferation, migration, and invasion. Overexpression of FOXA2 decreased, whereas knockout of FOXA2 increased VCAN mRNA and protein levels, which was due to direct binding and regulation of the VCAN gene by FOXA2. Interrogation of the TCGA ovarian cancer database revealed a negative relationship between FOXA2 and VCAN mRNA levels in EOC tumors, and high FOXA2/low VCAN mRNA levels in tumors positively correlated with patient survival. Finally, overexpression of FOXA2 or silencing of VCAN reversed the effects of mir-590. These findings demonstrate that miR-590-3p promotes EOC development via a novel FOXA2–VCAN pathway.Significance: Low FOXA2/high VCAN levels mediate the tumor-promoting effects of miR-590-3p and negatively correlate with ovarian cancer survival. Cancer Res; 78(15); 4175–90. ©2018 AACR.

Supplementary Figures 1-8 from The Ability of Versican to Simultaneously Cause Apoptotic Resistance and Sensitivity

Expression of the extracellular matrix proteoglycan versican is associated with more than 10 types of cancers, often being secreted by stromal cells in response to tumor signals. Previous work in our lab has shown that overexpression of the V1 versican isoform in cultured fibroblasts (V1 cells) increases both proliferation and apoptotic resistance. We show here that V1 cells induced tumor formation in nude mice and that, in keeping with previously shown apoptotic resistance, V1 cells have down-regulated Fas mRNA and protein levels. Unexpectedly, however, V1 cells were found to be sensitized to a wide range of cytotoxic agents. This combination of selective apoptotic resistance and sensitivity is often seen in cancer cells. V1 cells were also shown to have high resting levels of p53 and murine double minute-2 proteins, correlating with apoptotic sensitivity. Treatment with UV radiation induced p21 expression in vector-transfected cells but not in V1 cells. As p21 induces cell cycle arrest and inhibits apoptosis, its loss in V1 cells, coupled with high resting levels of proapoptotic p53, may be at least partially involved in their premature death following cytotoxic treatment. This study further supports the importance of versican in cancer cell biology and the complexity of apoptosis regulation. [Cancer Res 2007;67(10):4742–50]

Inflammation, a hallmark of cancer, has been associated with tumor progression, transition into malignant phenotype and efficacy of the chemotherapeutic agents in cancer. Chronic inflammation provides a favorable environment for tumorigenesis by inducing immunosuppression, whereas acute inflammation prompts tumor suppression by generating anti-tumor immune responses. Inflammatory factors derived from interstitial cells or tumor cells can stimulate cell proliferation and survival by modulating oncogenes and/or tumor suppressors. Recently, a new class of RNAs i.e., circular RNAs (circRNAs) has been implicated in inflammatory diseases. Although there are reports on circRNAs imparting functions in inflammatory insults, whether these circularized transcripts hold the potential to regulate inflammation induced cancer or tumor related inflammation, and modulate the interactions between tumor microenvironment (TME) and the inflammatory stromal/immune cells awaits further elucidation. Contextually, the current review describes the molecular association between inflammation and cancer, and spotlights the regulatory mechanisms by which circRNAs can moderate TME in response to inflammatory signals/ triggers. We also present comprehensive information about the immune cell(s)-specific expression and functions of the circRNAs in TME, modulation of inflammatory signaling pathways to drive tumorigenesis, and their plausible roles in inflammasomes and tumor development. Moreover, the diagnostic/prognostic and therapeutic potential of these circRNAs in harnessing inflammatory responses in cancer is also discussed.

Circular RNAs (circRNAs) are a large class of noncoding RNAs, generated from a process called back-splicing, that possess critical regulatory functions in many cellular events. A large body of literature has reported various circRNA functions and their underlying mechanisms, including sponging miRNA, exerting transcriptional and translational regulation, interacting with proteins, and translating into peptides and proteins. CircRNA dysregulation has been implicated in many cancers, including lung, breast, liver, gastric, colorectal, and ovarian cancer. They are detectable in bodily fluids and relatively stable, making them potential cancer biomarker candidates. Furthermore, targeting circRNA expression levels is a potential therapeutic approach for treating cancers. In this review, we describe the functional mechanisms of circRNAs and discuss limitations of current mechanism studies. Following this, we outline the potential of circRNAs to be effective biomarkers in various cancers and present circRNA-based therapeutic approaches. Finally, we discuss challenges in using circRNAs as diagnostic and therapeutic tools and propose future research directions.

Circular RNAs (circRNAs) represent a large group of non-coding RNAs that are widely detected in mammalian cells. Although most circRNAs are generated in a sense orientation, there is a group of circRNAs that are synthesized in an antisense orientation. High-throughput analysis of breast cancer specimens revealed a significant enrichment of 209 antisense circRNAs. The tumor suppressor SCRIB was shown to potentially produce thirteen circRNAs, three of which are in an antisense orientation. Among these three circRNAs, circSCRIB (hsa_circ_0001831) was the most enriched in the breast cancer panel. This antisense SCRIB circRNA was shown to span one intron and two exons. We hypothesized that this circRNA could decrease pre-mRNA splicing and mRNA translation. To test this, we generated a hsa_circ_0001831 expression construct. We found that there was decreased SCRIB mRNA production but increased cancer cell proliferation, migration, and invasion. In comparison, an exonic sequence construct did not affect mRNA splicing but decreased protein translation, leading to increased E-cadherin expression and decreased expression of N-cadherin and vimentin. Thus, there was increased cell migration, invasion, proliferation, colony formation, and tumorigenesis. Our study suggests a novel modulatory role of antisense circRNAs on their parental transcripts. This may represent a promising approach for developing circRNA-directed therapy.

Background: Fibrotic cardiac remodeling is a maladaptive response to acute or chronic injury that leads to arrhythmia and progressive heart failure. The underlying mechanisms remain unclear. We performed high-throughput RNA sequencing to analyze circular RNA profile in human cardiac disease and developed transgenic mice to explore the roles of circNlgn. Methods and Results: Using RNA sequencing, we found that circular neuroligin RNA (circNlgn) was highly upregulated in myocardial tissues of patients with selected congenital heart defects with cardiac overload. Back-splicing of the neuroligin gene led to the translation of a circular RNA–derived peptide (Nlgn173) with a 9-amino-acid nuclear localization motif. Binding of this motif to the structural protein LaminB1 facilitated the nuclear localization of Nlgn173. CHIP analysis demonstrated subsequent binding of Nlgn173 to both ING4 (inhibitor of growth protein 4) and C8orf44-SGK3 (serum and glucocorticoid-inducible kinase-3) promoters, resulting in aberrant collagen deposition, cardiac fibroblast proliferation, and reduced cardiomyocyte viability. Three-dimensional ultrasound imaging of circNlgn-transgenic mice showed impaired left ventricular function, with further impairment when subjected to left ventricular pressure overload compared with WT (wild type) mice. Nuclear translocation of Nlgn173, dysregulated expression of ING4 and C8orf44-SGK3, and immunohistochemical markers of cardiac fibrosis were detected in a panel of 145 patient specimens. Phenotypic changes observed in left ventricular pressure overload and transgenic mice were abrogated with silencing of circNlgn or its targets ING4 and SGK3. Conclusions: We show that a circular RNA can be translated into a novel protein isoform. Dysregulation of this process contributes to fibrosis and heart failure in cardiac overload–induced remodeling. This mechanism may hold therapeutic implications for cardiac disease.

Circular RNAs (circRNAs) are a group of non-coding RNAs with a unique circular structure generated by back-splicing. It is acknowledged that circRNAs play critical roles in cardiovascular diseases. However, functional studies of circRNAs were impeded due to lack of effective in vivo silencing approaches. Since most circRNAs are produced by protein-coding transcripts, gene editing typically affects the coding activity of the parental genes. In this study, we developed a circular antisense RNA (cA-circSlc8a1) that could silence the highly expressed circRNA circSlc8a1 in the mouse heart but not its parental Slc8a1 linear mRNA. Transgenic cA-circSlc8a1 mice developed congestive heart failure resulting in a significant increase in the body weight secondary to peripheral edema and congestive hepatopathy. To further test the role of circSlc8a1, we generated transgenic mice overexpressing circSlc8a1 and observed a protective effect of circSlc8a1 in a pressure overload model. Mechanistically, we found that circSlc8a1 translocated into mitochondria to drive ATP synthesis. While establishing a transgenic murine model for antisense-mediated circRNA silencing without interfering with the parental linear RNA, our finding revealed the essential role of circSlc8a1 in maintaining heart function and may lay the groundwork of using the circular antisense RNA as a potential gene therapy approach for cardiovascular diseases.

The Hippo signaling pathway is an evolutionarily conserved network that regulates organ size and tissue homeostasis in mammals. This pathway controls various cell functions, such as growth, proliferation, survival, apoptosis, and stemness by switching 'on' or 'off' its inhibitory and/or transcriptional module, thereby regulating target gene(s) expression. Altered Hippo signaling has been implicated in various forms of cancers. Increasing evidence suggests cross-talk between the Hippo signaling pathway and non-coding RNAs, in particular circular RNAs (circRNAs). In this context, the current review presents the mechanistic interplay between the Hippo pathway and related circRNAs in various forms of cancers, along with the capabilities of these circRNAs to function either as tumor suppressors or oncogenes through miRNA sponging or protein binding mechanisms. Furthermore, we discuss the constraints and limitations in circRNA mechanistic studies while highlighting some outstanding questions regarding the roles of circRNAs associated with the Hippo-YAP pathway in cancer. Finally, we delineate the potential of these circRNAs to be employed as diagnostic and prognostic biomarkers, as well as molecular hotspots for cancer therapy.

Circular RNAs (circRNAs) are circularized, single-stranded RNAs that are covalently linked. With their abundance in tissues and developmental stage-specific expression, circRNAs participate in a variety of physiological and pathological processes. In this review, we discuss the development of circRNAs used as biomarkers and therapeutic targets for cardiovascular diseases (CVDs), focusing on recent discoveries and applications of exosomal circRNAs that highlight opportunities and challenges. Some studies have identified a spectrum of circRNAs that are differentially expressed in CVDs, while other studies further manipulated specific circRNA expression and showed an ameliorated pathogenic state such as ischemic injury, hypertrophy, and cardiac fibrosis. Studies and applications of circRNAs are being rapidly developed. We expect to see clinical use of circRNAs as biomarkers and targets for disease treatment in the near future., Graphical Abstract, Large numbers of differentially expressed circRNAs have been identified in pathogenic heart tissues and cells. Gain-of-function and loss-of-function approaches are being developed to manipulate specific circRNA expression in order to modulate heart conditions. circRNAs may be used as potential biomarkers for early diagnosis and may be targets for therapeutic strategies.

BACKGROUND: Dietary polyphenols including anthocyanins target multiple organs. OBJECTIVE: We aimed to assess the involvement of glucagon-like peptide 1 (GLP-1), leptin, insulin and fibroblast growth factor 21 (FGF21) in mediating metabolic beneficial effects of purified anthocyanin cyanidin-3-glucoside (Cy3G). METHODS: Intestinal proglucagon gene (Gcg; encoding GLP-1) and liver Fgf21 expression were assessed in 6-wk-old male C57BL-6J mice fed a low-fat-diet (LFD; 10% of energy from fat), alone or with 1.6 mg Cy3G/L in drinking water for 3 wk [experiment (Exp.) 1; n = 5/group]. Similar mice were fed the LFD or a high-fat diet (HFD; 60% energy from fat) with or without Cy3G for 20 wk. Half of the mice administered Cy3G also received 4 broad-spectrum antibiotics (ABs) in drinking water between weeks 11 and 14, for a total of 6 groups (n = 8/group). Metabolic tolerance tests were conducted between weeks 2 and 16. Relevant hormone gene expression and plasma hormone concentrations were assessed mainly at the end of 20 wk (Exp. 2). RESULTS: In Exp. 1, Cy3G administration increased ileal but not colonic Gcg level by 2-fold (P 3-fold, P

Circular RNAs (circRNAs) are ubiquitously expressed, covalently closed rings, produced by pre-mRNA splicing in a reversed order during post-transcriptional processing. Circularity endows 3'-5'-linked circRNAs with stability and resistance to exonucleolytic degradation which raises the question whether circRNAs may be relevant as potential therapeutic targets or agents. High stability in biological systems is the most remarkable property and a major criterion for why circRNAs could be exploited for a range of RNA-centred medical applications. Even though various biological roles and regulatory functions of circRNAs have been reported, their in-depth study is challenging because of their circular structure and sequence-overlap with linear mRNA counterparts. Moreover, little is known about their role in viral infections and in antiviral immune responses. We believe that an in-depth and detailed understanding of circRNA mediated viral protein regulations will increase our knowledge of the biology of these novel molecules. In this review, we aimed to provide a comprehensive basis and overview on the biogenesis, significance and regulatory roles of circRNAs in the context of antiviral immune responses and viral infections including hepatitis C virus infection, hepatitis B virus infection, hepatitis delta virus infection, influenza A virus infection, Epstein-Barr virus infection, kaposi's sarcoma herpesvirus infection, human cytomegalovirus infection, herpes simplex virus infection, human immunodeficiency virus infection, porcine epidemic diarrhoea virus infection, ORF virus infection, avian leukosis virus infection, simian vacuolating virus 40 infection, transmissible gastroenteritis coronavirus infection, and bovine viral diarrhoea virus infection. We have also discussed the critical regulatory role of circRNAs in provoking antiviral immunity, providing evidence for implications as therapeutic agents and as diagnostic markers.

The mushroom Ganoderma lucidum (G. lucidum Leyss. ex Fr.) Karst has been a traditional Chinese medicine for millennia. In this study, we isolated the Ganoderma lucidum spore oil (GLSO) and evaluated the effect of GLSO on skin burn wound healing and the underlying mechanisms. Mice were used to perform skin wound healing assay. Wound analysis was performed by photography, hematoxylin/eosin staining, Masson's Trichrome staining and immunohistochemical analysis. Microbiota on the wounds were analyzed using the 16s rRNA sequence and quantitative statistics. The lipopolysaccharide (LPS) content was examined in skin wounds and serum using an enzyme-linked immunosorbent assay (ELISA). The expression of Toll-like receptor 4 (TLR4) and the relative levels of inflammatory cytokines were determined by qPCR and immunofluorescence assay. A pseudo-germfree mouse model treated with antibiotics was used to investigate whether GLSO accelerated skin burn wound healing through the skin microbiota. We found that GLSO significantly accelerated the process of skin wound healing and regulated the levels of gram-negative and gram-positive bacteria. Furthermore, GLSO reduced LPS and TLR4, and levels of some other related inflammatory cytokines. The assay with the pseudo-germfree mice model showed that GLSO had a significant acceleration on skin wound healing in comparison with antibiotic treatment. Thus, GLSO downregulated the inflammation by regulating skin microbiota to accelerate skin wound healing. These findings provide a scientific rationale for the potential therapeutic use of GLSO in skin burn injury.

Metastatic cancer cells invade surrounding tissues by forming dynamic actin-based invadopodia, which degrade the surrounding extracellular matrix and allow cancer cell invasion. Regulatory RNAs, including circular RNA, have been implicated in this process. By microarray, we found that the circular RNA circSKA3 was highly expressed in breast cancer cells and human breast cancer tissues. We further found that the invasive capacity of breast cancer cells was positively correlated with circSKA3 expression, through the formation of invadopodia. Mechanistically, we identified Tks5 and integrin β1 as circSKA3 binding partners in these tumor-derived invadopodia. Ectopic circSKA3 expression conferred increased tumor invasiveness in vitro and in vivo. We further identified the RNA-protein binding sites between circSKA3, Tks5 and integrin β1. In tumor formation assays, we found that circSKA3 expression promoted tumor progression and invadopodium formation. Mutation of the circSKA3 binding sites or transfection with blocking oligos abrogated the observed effects. Thus, we provide evidence that the circular RNA circSKA3 promotes tumor progression by complexing with Tks5 and integrin β1, inducing invadopodium formation.

Cynomorium coccineum has long been used as the health and medicinal plant known to induce cancer cell death. However, the bioactive compounds of C. coccineum and the underlying mechanism of their regulator in cell autophagy and cell apoptosis remain unexplored. In our previous study, we found that the ethanol extract had antitumor activity through inducing cancer cell death. In this study, by detecting the anti-tumor effect of sequence extracts from Cynomorium coccineum, the active constituents were collected in solvent ethyl acetate. A strategy based on ultra-performance liquid chromatography coupled with hybrid quadrupole-orbitrap mass spectrometry (UPLC-Q-Orbitrap/MS) was first utilized to analyze the chemical constituents of active fraction (ethyl acetate fraction, CS3). A total of 29 compounds including 8 triterpenoids, 6 flavonoids, 4 fatty acids, 8 phenolic acids, 1 anthraquinones, 1 nucleoside and 1 sterol were detected and identified or tentatively identified for the first time in Cynomorium coccineum. We found that CS3 induces cancer cell death accompanied with a great number of vacuoles in the cytoplasm. CS3-induced autophagosome formation was found and confirmed by electron microscopy and the high expression levels of microtubule-associated protein-1 light chain 3-II (LC3II), a marker protein of autophagy. We additionally demonstrated that CS3 activated and increased the pro-apoptotic mitochondrial proteins, BNIP3 and BNIP3L, in mRNA and protein levels. The constituents of CS3 down-regulated anti-apoptotic BCL2, and then releases autophagic protein Beclin-1. These finding for the first time systematically not only explore and identify the active constituents of CS3 in Cynomorium coccineum, but also examined the mechanism associated with CS3-induced cell death via cell autophagy. This active component may serve as a potential source to obtain new autophagy inducer and anti-cancer compounds for hepatocellular carcinoma.

Our pre-experiments found that the brain circRNA sequence profiles and gut microbiota in AD-like mice were changed, as circNF1-419 could enhance autophagy to ameliorate senile dementia in AD-like mice, so we conclude that there might some connections between circRNA and gut microbiome. Therefore, we use the over-expressed circNF1-419 adeno-associated virus (AAV) animal system with the aim of identifying possible connections. Our results showed that over-expression of circNF1-419 in brain not only influenced the cholinergic system of brain, but also changed the gut microbiota composition as the Candidatus Arthromitus, Lachnospiraceae FCS020 group, Lachnospiraceae UCG-006, and [Eubacterium] xylanophilum group, and the intestinal homeostasis and physiology, and even the gut microbiota trajectory in new born mice. These findings demonstrate a link between circRNA and gut microbiome, enlarge the 'microbiome- transcriptome' linkage library and provide more information on gut-brain axis.

Recent studies have demonstrated circular RNAs (circRNAs) to be widely expressed and to have important physiological functions. However, the expression, regulation, and function of circRNAs in neuroglial cells are unknown. Herein, we characterized the expression, regulation, and function of circRNAs in astrocytes. Astrocyte circRNAs were identified by computational analysis of newborn SD rat primary astrocytes cultured with 20 g/L D-galactose. In this manner, 7376 circRNAs were identified, among which most circRNAs (5754) were derived from annot_exons, whereas 27 were antisense, 853 were exon/intron, 329 were intergenic, 41 were intronic, and 372 were one exon. Among these, circNF1-419 was demonstrated to regulate autophagy, in over-expressing circNF1-419 transfected astrocytes, through the PI3K-I/Akt-AMPK-mTOR and PI3K-I/Akt-mTOR signaling pathways. An adenovirus associated virus packaging system (virus titer 1 ×1012), over-expressing circNF1-419 and injected into mouse cerebral cortex, showed autophagy enhancing activity by binding the proteins Dynamin-1 and Adaptor protein 2 B1 (AP2B1). This binding regulated aging markers (p21, p35/25, and p16) and inflammatory factors (TNF-α and NF-κB), and reduced the expression of Alzheimer’s disease marker proteins (Tau, p-Tau, Aβ1-42, and APOE), which delayed senile dementia. Transcriptome analysis of the brain showed that circNF1-419 improved other signaling pathways, especially those related to the synapses of SAMP8 mice. These findings provide novel insights into circNF1-419 and its potential usefulness for the diagnosis and treatment of dementia by regulating Dynamin-1 and AP2B1 mediated autophagy.

Chemoresistance of triple negative breast cancer against paclitaxel (PAX) is one of the major issues for the patients under chemotherapy. However, the mechanism by which the breast cancer cells are resistant to PAX remains unclear. Here, we identified a circular RNA of angiomotin-like 1 (circAMOTL1) as an important player which may be responsible for the adverse resistance against PAX in breast cancer cells. The circAMOTL1 were overexpressed in MDA-MB-231 breast cancer cells via transfection of circAMOTL1 construct. Overexpression of circAMOTL1 caused significant increase of cell viability, reduction of apoptosis, and enhancement of invasion when MDA-MB-231 cells were exposed to PAX compared to those cells with vector control. Moreover, these resistant effects could be blocked by the application of circAMOTL1 siRNA. In these circAMOTL1 overexpressing cells, we found notable increase of both phosphorylated and total AKT protein, which suggested that AKT might be the downstream factor mediating the resistant effects. Consequently, the gene and protein expression of AKT related pro-apoptotic (BAX and BAK) and anti-apoptotic (BCL-2) factors were significantly changed by circAMOTL1 as well. These results suggest circAMOTL1 may play an important role in the PAX resistance of breast cancer cells via regulation of AKT pathway, facilitation of anti-apoptotic protein and inhibition of pro-apoptotic protein. While providing a new mechanism of PAX resistance in breast cancer cells, our findings may lay groundwork for a novel therapeutic target of the breast cancer treatment in the future.

In recent years, circular RNAs (circRNAs), a new class of RNA molecules characterized by their covalently closed circular structure, have become a new research paradigm in RNA biology. Many circRNAs are conserved among eukaryotes, localize in specific subcellular compartments, and play different biological roles. Accumulating evidence shows that circRNAs regulate a diversity of cellular processes by acting as miRNA sponges, anchors for circRNA binding proteins (cRBPs), transcriptional regulators, molecular scaffolds, and sources for translation of small proteins/peptides. The emergence of the biological functions of circRNAs has brought a new perspective to our understanding of cellular physiology and disease pathogenesis. Recent studies have shown that the expression of circRNAs is tissue- and cell type-specific and specifically regulated through development or disease progression, where they exert specific biological functions. However, the mechanisms underlying these remain largely unknown. A deeper understanding of how the specific expression of circRNAs is regulated to exert specific biological functions will enable the use of circRNA as a biomarker in clinical practice and the development of new therapeutic approaches. This review aims to summarize recent developments in circRNA biogenesis, functions, and molecular mechanisms. We also provide some specific circRNAs as examples to show their tissue-specific distribution and evaluate the possibility of applying circRNA technologies in molecular research and therapeutics.