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A systematic mammalian genetic interaction map reveals pathways underlying ricin susceptibility.
Publication Type:Journal Article
Source:Cell, Volume 152, Issue 4, p.909-22 (2013)
Keywords:Biological Transport, Carrier Proteins, Cell Line, Tumor, Coat Protein Complex I, Endoplasmic Reticulum, Epistasis, Genetic, Heptanoic Acids, Humans, Membrane Proteins, Proto-Oncogene Proteins, Pyrroles, Ribosomal Proteins, Ricin, RNA, Small Interfering, Vesicular Transport Proteins
<p>Genetic interaction (GI) maps, comprising pairwise measures of how strongly the function of one gene depends on the presence of a second, have enabled the systematic exploration of gene function in microorganisms. Here, we present a two-stage strategy to construct high-density GI maps in mammalian cells. First, we use ultracomplex pooled shRNA libraries (25 shRNAs/gene) to identify high-confidence hit genes for a given phenotype and effective shRNAs. We then construct double-shRNA libraries from these to systematically measure GIs between hits. A GI map focused on ricin susceptibility broadly recapitulates known pathways and provides many unexpected insights. These include a noncanonical role for COPI, a previously uncharacterized protein complex affecting toxin clearance, a specialized role for the ribosomal protein RPS25, and functionally distinct mammalian TRAPP complexes. The ability to rapidly generate mammalian GI maps provides a potentially transformative tool for defining gene function and designing combination therapies based on synergistic pairs.</p>