KaryoCreate: A CRISPR-based technology to study chromosome-specific aneuploidy by targeting human centromeres

Aneuploidy, the presence of chromosome gains or losses, is a hallmark of cancer. Here, we describe KaryoCreate (karyotype CRISPR-engineered aneuploidy technology), a system that enables the generation of chromosome-specific aneuploidies by co-expression of an sgRNA targeting chromosome-specific CENPA-binding ɑ-satellite repeats together with dCas9 fused to mutant KNL1. We design unique and highly specific sgRNAs for 19 of the 24 chromosomes. Expression of these constructs leads to missegregation and induction of gains or losses of the targeted chromosome in cellular progeny, with an average efficiency of 8% for gains and 12% for losses (up to 20%) validated across 10 chromosomes. Using KaryoCreate in colon epithelial cells, we show that chromosome 18q loss, frequent in gastrointestinal cancers, promotes resistance to TGF-β, likely due to synergistic hemizygous deletion of multiple genes. Altogether, we describe an innovative technology to create and study chromosome missegregation and aneuploidy in the context of cancer and beyond. [Display omitted] •We designed chromosome-specific sgRNAs for targeting 19 of 24 human chromosomes•KaryoCreate facilitates sgRNA-dCas9 mediated recruitment of mutant kinetochore proteins•KaryoCreate enables the creation of human cells with distinct karyotypes•Engineered 18q loss promotes tumor-associated phenotypes in colon epithelial cells A method for engineering human cells with distinct karyotypes of interest enables interrogation of a specific aneuploidy in the context of cancer.