Graduation Year


Document Type




Degree Name

Master of Science (M.S.)

Degree Granting Department

Biology (Cell Biology, Microbiology, Molecular Biology)

Major Professor

Margaret Park, Ph.D.

Co-Major Professor

Charles Chalfant, Ph.D.

Committee Member

Sandy Westerheide, Ph.D.

Committee Member

Florian Karreth, Ph.D.


alternative splicing, bioinformatics, circular RNA, proliferation, triple negative breast cancer


Triple-negative breast cancer (TNBC) comprises only 24% of breast cancer cases, yet is the second leading cause of cancer mortality in women due to its aggressive nature (1). This increase in mortality is due to the lack of receptors for three targetable growth factors (HER2, progesterone, and estrogen receptors). Our previous studies have indicated that these cancers are highly dysregulated in respect to alternative splicing. Hence, we undertook a study aimed at identifying circular RNAs (circRNAs) generated from back-splicing events which were dysregulated in TNBC. We have identified a novel circRNA transcript, circular REV1 (circREV1), which is upregulated in our TNBC cell lines. Its overexpression may be an indicator of TNBC and its progression. The complexes formed between circRNAs and proteins or other RNA transcripts are able to dysregulate gene expression, which is one of the hallmarks of cancer (2).Next generation sequencing of RNA collected from breast epithelium and TNBC cell lines were aligned with STAR aligner and bioinformatically analyzed for differential expression of circRNAs via circtools (3). CircREV1 was found to be overexpressed in two TNBC cell lines, while demonstrating minimal expression in breast epithelial cells. Quantitative polymerase chain reaction (qRT-PCR) analysis, following reverse transcription of RNA collected from relevant cell lines, validated these findings in vitro. Treatment with RNase R, to remove the linear construct of REV1, confirmed that the circular isoform is responsible for the overexpression found in the TNBC cell lines. The isoform we have honed in on is that of exon 3 back-spliced to exon 2, with the intron spanning the two being retained. Linear REV1 is suggested to function as a scaffolding protein to recruit DNA polymerases for translesion synthesis in DNA repair mechanisms (4). It can be hypothesized that cancer may manipulate this gene to facilitate bypass of cell cycle regulation machinery. While minimal data has been published on the function of circREV1, it is predicted to be involved in a pathway which favors cancer advancement. Ultimately, our findings suggest that circREV1 upregulation may be influential in the transformation from breast epithelium to a more aggressive phenotype.