Most RNAs generated by the human genome have no protein-coding ability and are termed non-coding RNAs. Among these include circular RNAs, which include exonic circular RNAs (circRNA), mainly found in the cytoplasm, and intronic RNAs (ciRNA), predominantly detected in the nucleus. The biological functions of circular RNAs remain largely unknown, although ciRNAs have been reported to promote gene transcription, while circRNAs may function as microRNA sponges. We demonstrate that the circular RNA circ-Foxo3 was highly expressed in non-cancer cells and were associated with cell cycle progression. Silencing endogenous circ-Foxo3 promoted cell proliferation. Ectopic expression of circ-Foxo3 repressed cell cycle progression by binding to the cell cycle proteins cyclin-dependent kinase 2 (also known as cell division protein kinase 2 or CDK2) and cyclin-dependent kinase inhibitor 1 (or p21), resulting in the formation of a ternary complex. Normally, CDK2 interacts with cyclin A and cyclin E to facilitate cell cycle entry, while p21works to inhibit these interactions and arrest cell cycle progression. The formation of this circ-Foxo3-p21-CDK2 ternary complex arrested the function of CDK2 and blocked cell cycle progression.

MicroRNAs (miRNAs) are a class of 20–24 nt non-coding RNAs that regulate gene expression primarily through post-transcriptional repression or mRNA degradation in a sequence-specific manner. The roles of miRNAs are just beginning to be understood, but the study of miRNA function has been limited by poor understanding of the general principles of gene regulation by miRNAs. Here we used CNE cells from a human nasopharyngeal carcinoma cell line as a cellular system to investigate miRNA-directed regulation of VEGF and other angiogenic factors under hypoxia, and to explore the principles of gene regulation by miRNAs. Through computational analysis, 96 miRNAs were predicted as putative regulators of VEGF. But when we analyzed the miRNA expression profile of CNE and four other VEGF-expressing cell lines, we found that only some of these miRNAs could be involved in VEGF regulation, and that VEGF may be regulated by different miRNAs that were differentially chosen from 96 putative regulatory miRNAs of VEGF in different cells. Some of these miRNAs also co-regulate other angiogenic factors (differential regulation and co-regulation principle). We also found that VEGF was regulated by multiple miRNAs using different combinations, including both coordinate and competitive interactions. The coordinate principle states that miRNAs with independent binding sites in a gene can produce coordinate action to increase the repressive effect of miRNAs on this gene. By contrast, the competitive principle states when multiple miRNAs compete with each other for a common binding site, or when a functional miRNA competes with a false positive miRNA for the same binding site, the repressive effects of miRNAs may be decreased. Through the competitive principle, false positive miRNAs, which cannot directly repress gene expression, can sometimes play a role in miRNA-mediated gene regulation. The competitive principle, differential regulation, multi-miRNA binding sites, and false positive miRNAs might be useful strategies in the avoidance of unwanted cross-action among genes targeted by miRNAs with multiple targets.

Circular RNAs have been identified as naturally occurring RNAs that are highly represented in the eukaryotic transcriptome. Although a large number of circRNAs have been reported, circRNA functions remain largely unknown. CircRNAs can function as miRNA sponges, thereby reducing their ability to target mRNAs. We hypothesize that circRNAs may bind, store, sort, and sequester proteins to particular subcellular locations, and act as dynamic scaffolding molecules that modulate protein-protein interactions. Here, we review the biological implication and function of circRNA-protein interaction, and reveal a dynamic model of the interaction in various tissues, development stages and physiological conditions. Improved techniques to identify and characterize the dynamic RNA-protein interactions may elucidate the molecular mechanisms associated with the expression and functional diversity of circRNAs.

Significant progress has been made in circular RNA (circRNA) research in recent years. Increasing evidence suggests that circRNAs play important roles in many cellular processes, and their dysregulation is implicated in the pathogenesis of various diseases. CircRNAs are highly stable and usually expressed in a tissue- or cell type-specific manner. Therefore, they are currently being explored as potential therapeutic targets. Gain-of-function and loss-of-function approaches are typically performed using circRNA expression plasmids and RNA interference-based strategies, respectively. These strategies have limitations that can be mitigated using nanoparticle and exosome delivery systems. Furthermore, recent developments show that the cre-lox system can be used to knockdown circRNAs in a cell-specific manner. While still in the early stages of development, the CRISPR/Cas13 system has shown promise in knocking down circRNAs with high specificity and efficiency. In this review, we describe circRNA properties and functions and highlight their significance in disease. We summarize strategies that can be used to overexpress or knockdown circRNAs as a therapeutic approach. Lastly, we discuss major challenges and propose future directions for the development of circRNA-based therapeutics.

Circular RNAs (circRNAs) represent a new type of regulatory RNA which forms a covalently closed continuous loop from back-splicing events, a process in which the downstream 5′ splice site and the 3′ splice site are covalently linked. Emerging evidence indicates that circRNAs exert a new layer of transcriptional and post-transcriptional regulation of gene expression. However, there is no standard nomenclature of circRNA, although the study of circRNAs has exploded in the past few years. Here we present circbank (www.circbank.cn), a comprehensive database for human circRNAs, where a novel naming system of circRNAs based on the host genes of circRNAs was implemented. In addition to the new naming system, circbank collected other five features of circRNAs including the miRNA binding site, conservation of circRNAs, m⁶A modification of circRNAs, mutation of circRNAs and protein-coding potential of circRNAs. Circbank is publicly available and allows users to query, browse and download circRNAs with all six features we provided, based on different search criteria. The database may serve as a resource to facilitate the research of function and regulation of circRNAs.