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

Yongchao Dou,James G Patton,Bing Zhang,Alissa M Weaver,Diana J Cha,Shawn Levy,Jeffrey L Franklin,Dennis K Jeppesen,Robert J Coffey,James N Higginbotham,Nripesh Prasad

doi:10.1038/srep37982

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.

Highlights

Knowledge about the general sequence features, biogenesis, and putative functions of circRNAs, especially exonic circRNAs, has gradually accumulated[11]
We showed that circRNA abundance was down-regulated at a global level in mutant KRAS cell lines, suggesting a potential involvement of circRNAs in oncogenesis
One possibility is that down-regulation of circRNAs in KRAS mutant cells is caused by their increased exporting to exosomes

Summary

Introduction

Knowledge about the general sequence features, biogenesis, and putative functions of circRNAs, especially exonic circRNAs, has gradually accumulated[11] Because both circRNAs and linear RNAs are spliced from pre-mRNAs, the competition between circularization and linear splicing may play a role in the regulation of gene www.nature.com/scientificreports/. We studied both cellular and exosomal circRNAs in the three cell lines, with confirmation of altered circRNAs in a second set of isogenically matched cell lines. To our knowledge, this is the first report describing the impact of a well-established oncogene on the abundance of circRNAs

Methods

Results

Conclusion