APOBEC3B reporter myeloma cell lines identify DNA damage response pathways leading to APOBEC3B expression

Abstract

Apolipoprotein B mRNA-editing enzyme catalytic polypeptide-like (APOBEC) DNA cytosine deaminase 3B (A3B) is a DNA editing enzyme which induces genomic DNA mutations in multiple myeloma and various other cancers. APOBEC family proteins are highly homologous so it is especially difficult to investigate the biology of A3B alone in cancer cells. To investigate A3B function in myeloma cells easily and comprehensively, we used CRISPR/Cas9 to generate A3B reporter cells that contain 3×FLAG tag and IRES-EGFP sequences integrated at the end of the A3B gene. These reporter cells stably express 3xFLAG tagged A3B and the reporter EGFP and this expression is enhanced under known stimuli, such as PMA. Conversely, shRNA knockdown of A3B decreased EGFP fluorescence and 3xFLAG tagged A3B protein levels. We screened a series of anticancer treatments using these cell lines and identified that most conventional therapies, such as antimetabolites or radiation, exacerbated endogenous A3B expression, but recent molecular targeting drugs, including bortezomib, lenalidomide and elotuzumab, did not. Furthermore, chemical inhibition of ATM, ATR and DNA-PK suppressed the EGFP expression upon treatment with antimetabolites. These results suggest that DNA damage response triggers A3B expression through ATM, ATR and DNA-PK signaling.