Induction of t(11;14) IgH enhancer/promoter- gene translocation using CRISPR/Cas9.

Chromosomal translocation is a key process in the oncogenic transformation of somatic cells. Previously, artificial induction of chromosomal translocation was performed using homologous recombination-mediated loxP labeling of target regions followed by Cre-mediated recombination. Recent progress in genome editing techniques has facilitated the easier induction of artificial translocation by cutting two targeted genome sequences from different chromosomes. The present study established a system to induce t(11;14)(q13;q32), which is observed primarily in multiple myeloma (MM) and involves the repositioning of the () gene downstream of the immunoglobulin heavy chain (IgH) constant region enhancers by translocation. The placing of tandem gRNAs designed to cut both the and 15-kb upstream regions in lentiCRISPRv2 enabled the induction of chromosomal translocation in 293T cells, with confirmation by translocation-specific PCR and fluorescence hybridization probing with and . At the translocation junctions, small deletions and the addition of DNA sequences (indels) were observed in several clones. Cloned cells with t(11;14) exhibited slower growth and lower mRNA expression compared to the parent cells, presenting the opposite phenomena induced by t(11;14) in MM cells, indicating that the silent gene juxtaposed to may negatively affect gene expression and cell proliferation in the non-B lymphocyte lineage. Therefore, the present study achieved the induction of silent promoter/enhancer translocation in t(11;14)(q13;q32) as a preparatory experiment to study the role of constant region enhancer-driven overexpression in oncogenic transformation processes in B lymphocytes.