Circular RNAs are down-regulated in KRAS mutant colon cancer cells and can be transferred to exosomes

Abstract

Recent studies have shown that circular RNAs (circRNAs) are abundant, widely expressed in mammals, and can display cell-type specific expression. However, how production of circRNAs is regulated and their precise biological function remains largely unknown. To study how circRNAs might be regulated during colorectal cancer progression, we used three isogenic colon cancer cell lines that differ only in KRAS mutation status. Cellular RNAs from the parental DLD-1 cells that contain both wild-type and G13D mutant KRAS alleles and isogenically-matched derivative cell lines, DKO-1 (mutant KRAS allele only) and DKs-8 (wild-type KRAS allele only) were analyzed using RNA-Seq. We developed a bioinformatics pipeline to identify and evaluate circRNA candidates from RNA-Seq data. Hundreds of high-quality circRNA candidates were identified in each cell line. Remarkably, circRNAs were significantly down-regulated at a global level in DLD-1 and DKO-1 cells compared to DKs-8 cells, indicating a widespread effect of mutant KRAS on circRNA abundance. This finding was confirmed in two independent colon cancer cell lines HCT116 (KRAS mutant) and HKe3 (KRAS WT). In all three cell lines, circRNAs were also found in secreted extracellular-vesicles, and circRNAs were more abundant in exosomes than cells. Our results suggest that circRNAs may serve as promising cancer biomarkers.

Diverse Long RNAs Are Differentially Sorted into Extracellular Vesicles Secreted by Colorectal Cancer Cells

“…However, the diversity of exRNA is extensive and microRNAs (miRNAs) are not the most abundant class of RNA found in EVs (Fritz et al, 2016; Mateescu et al, 2017). Analysis of cellular versus exRNA has repeatedly demonstrated selective biogenesis, export, and/or stability of specific RNAs (Cha et al, 2015; Dou et al, 2016; Kosaka et al, 2010; Santangelo et al, 2016; Skog et al, 2008; Squadrito et al, 2014; Valadi et al, 2007; Villarroya-Beltri et al, 2013; Wei et al, 2017). Elucidation of the mechanisms for selective sorting of cargo into EVs is critical to understanding extracellular signaling by RNA.…”

“…We also found that specific intracellular oncogenic signaling events can regulate trafficking of miRNAs through phosphorylation of Argonaute (AGO) proteins (McKenzie et al, 2016). More recently, we identified a global downregulation of circular RNAs (circRNAs) in mutant KRAS cells with an inverse upregulation in EVs (Dou et al, 2016). Here, we report comprehensive analysis of EV long RNAs (>200 nt) and show that similar to miRNA export, there is selective export of long RNAs to EVs.…”

A Meta-Analysis of the Diagnostic Accuracy of Circular RNAs in Digestive System Malignancy

“…Fourth, most of the investigated circRNAs were extracted from tissue samples; only one study combined tissue and plasma to assess the diagnostic value of circRNAs [32]. As previously reported, circRNAs are enriched and stable in exosomes [51, 52], which indicates that it might be possible to diagnose cancer by detecting plasma or serum circRNAs. It is believed that circRNAs could be useful as a noninvasive tumor screening tool for clinical practice in the future [53].…”

Circulating Circles Predict Postoperative Atrial Fibrillation

“…Moreover, aberrant expression of numerous circRNAs in disease, and the detection of these RNA species in the bloodstream have raised the exciting possibility that they may be used as noninvasive biomarkers. Indeed, the first studies revealed that circRNAs can be secreted in exosomes10 and that circulating circRNAs may be used as diagnostic biomarkers for type 2 diabetes mellitus,11 for risk stratification after myocardial infarction,12 and for cancer diagnosis 13…”