KaryoCreate: a new CRISPR-based technology to generate chromosome-specific aneuploidy by targeting human centromeres

Aneuploidy, the presence of chromosome gains or losses, is a hallmark of cancer and congenital syndromes. Here, we describe KaryoCreate (Karyotype CRISPR Engineered Aneuploidy Technology), a system that enables generation of chromosome-specific aneuploidies by co-expression of a sgRNA targeting chromosome-specific CENPA-binding [a]-satellite repeats together with dCas9 fused to a mutant form of KNL1. We designed unique and highly specific sgRNAs for 19 out of 24 chromosomes. Expression of these sgRNAs with KNL1Mut-dCas9 leads to missegregation and induction of gains or losses of the targeted chromosome in cellular progeny with an average efficiency of 8% and 12% for gains and losses, respectively (up to 20%), tested and validated across 9 chromosomes. Using KaryoCreate in colon epithelial cells, we show that chromosome 18q loss, a frequent occurrence in gastrointestinal cancers, promotes resistance to TGF{beta}, likely due to synergistic hemizygous deletion of multiple genes. Altogether, we describe a novel technology to create and study chromosome missegregation and aneuploidy in the context of cancer and beyond. HighlightsO_LIWe designed sgRNAs targeting chromosome-specific centromeres across 19 human chromosomes C_LIO_LIKaryoCreate combines chromosome-specific centromeric sgRNAs with dCas9 fused to a mutant form of KNL1. C_LIO_LIKaryoCreate allows engineering gains and losses of specific human chromosomes. C_LIO_LIEngineered Chromosome 18q loss promotes tumor-associated phenotypes in colon-derived cells. C_LIO_LIKaryoCreate is a CRISPR-based technology to foster the study of centromere biology and aneuploidy. C_LI