by
Michael Boettcher;
Ruilin Tian;
James A. Blau;
Evan Markegard;
Ryan T. Wagner;
David Wu;
Xiulei Mo;
Anne Biton;
Noah Zaitlen;
Haian Fu;
Frank McCormick;
Martin Kampmann;
Michael T. McManus
Understanding the direction of information flow is essential for characterizing how genetic networks affect phenotypes. However, methods to find genetic interactions largely fail to reveal directional dependencies. We combine two orthogonal Cas9 proteins from Streptococcus pyogenes and Staphylococcus aureus to carry out a dual screen in which one gene is activated while a second gene is deleted in the same cell. We analyze the quantitative effects of activation and knockout to calculate genetic interaction and directionality scores for each gene pair. Based on the results from over 100,000 perturbed gene pairs, we reconstruct a directional dependency network for human K562 leukemia cells and demonstrate how our approach allows the determination of directionality in activating genetic interactions. Our interaction network connects previously uncharacterized genes to well-studied pathways and identifies targets relevant for therapeutic intervention.
by
Caitlin L. Grzeskowiak;
Samrat T. Kundu;
Xiulei Mo;
Andrey Andreyevich Ivanov;
Oksana Zagorodna;
Hengyu Lu;
Richard H. Chapple;
Yiu Huen Tsang;
Daniela Moreno;
Maribel Mosqueda;
Karina Eterovic;
Jared J. Fradette;
Sumreen Ahmad;
Femgju Chen;
Zechen Chong;
Ken Chen;
Chad J. Creighton;
Haian Fu;
Gordan B. Mills;
Don L. Gibbons;
Kenneth L. Scott
Genetic aberrations driving pro-oncogenic and pro-metastatic activity remain an elusive target in the quest of precision oncology. To identify such drivers, we use an animal model of KRAS-mutant lung adenocarcinoma to perform an in vivo functional screen of 217 genetic aberrations selected from lung cancer genomics datasets. We identify 28 genes whose expression promoted tumor metastasis to the lung in mice. We employ two tools for examining the KRAS-dependence of genes identified from our screen: 1) a human lung cell model containing a regulatable mutant KRAS allele and 2) a lentiviral system permitting co-expression of DNA-barcoded cDNAs with Cre recombinase to activate a mutant KRAS allele in the lungs of mice. Mechanistic evaluation of one gene, GATAD2B, illuminates its role as a dual activity gene, promoting both pro-tumorigenic and pro-metastatic activities in KRAS-mutant lung cancer through interaction with c-MYC and hyperactivation of the c-MYC pathway.