Image-based identification and isolation of micronucleated cells to dissect cellular consequences

Recent advances in isolating cells based on visual phenotypes have transformed our ability to identify the mechanisms and consequences of complex traits. Micronucleus (MN) formation is a frequent outcome of genome instability, triggers extensive disease-associated changes in genome structure and signaling coincident with MN rupture, and is almost exclusively defined by visual analysis. Automated MN detection in microscopy images has proved extremely challenging, limiting unbiased discovery of the mechanisms and consequences of MN formation and rupture. In this study we describe two new MN segmentation modules: a rapid and precise model for classifying micronucleated cells and their rupture status (VCS MN), and a robust model for accurate MN segmentation (MNFinder) from a broad range of microscopy images. As a proof-of-concept, we define the transcriptome of non-transformed human cells with intact or ruptured MN after inducing chromosome missegregation by combining VCS MN with photoactivation-based cell isolation and RNASeq. Surprisingly, we find that neither MN formation nor rupture triggers a unique transcriptional response. Instead, transcriptional changes are correlated with increased aneuploidy in these cell classes. Our MN segmentation modules overcome a significant challenge to reproducible MN quantification, and, joined with visual cell sorting, enable the application of powerful functional genomics assays, including pooled CRISPR screens and time-resolved analyses of cellular and genetic consequences, to a wide-range of questions in MN biology.