Genome-wide RNAi Screens Identify KRAS-NF-κB Signaling

Large-scale, arrayed format RNAi screens to identify genes essential for proliferation/viability were performed as described3 (link),14 (link). The effect of introducing each of the 5002 shRNAs (targeting 957 genes) was determined in 19 cell lines, and normalized using the B-score metric4 (link). Feature selection of shRNA B-score data was performed using the Comparative Marker Application Suite in GenePattern5 (link) and was independently analyzed using RIGER analysis6 (link) to compute NES for each gene. Secondary screen viability data was normalized using a percent of control statistic, given the biased nature of the candidate shRNA plate. Expression profiling was used to generate a signature that correlates with KRAS activation and implicated NF-κB signaling in cell lines and tumors dependent on KRAS. Regulation of NF-κB by TBK1 was shown using biochemical and cell biological approaches. Details of the analytical methods are provided in the Full Methods.

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Barbie D.A., Tamayo P., Boehm J.S., Kim S.Y., Moody S.E., Dunn I.F., Schinzel A.C., Sandy P., Meylan E., Scholl C., Fröhling S., Chan E.M., Sos M.L., Michel K., Mermel C., Silver S.J., Weir B.A., Reiling J.H., Sheng Q., Gupta P.B., Wadlow R.C., Le H., Hoersch S., Wittner B.S., Ramaswamy S., Livingston D.M., Sabatini D.M., Meyerson M., Thomas R.K., Lander E.S., Mesirov J.P., Root D.E., Gilliland D.G., Jacks T, & Hahn W.C. (2009). Systematic RNA interference reveals that oncogenic KRAS-driven cancers require TBK1. Nature, 462(7269), 108-112.

Publication 2009

Biological Cell Cell lines Gene Kras Nf κb Rnai Shrna Tbk1 Tumors

Corresponding Organization :

Other organizations : Broad Institute, Massachusetts Institute of Technology, Brigham and Women's Hospital, Harvard University, Max Planck Institute for Metabolism Research, Max Planck Society, Allen Institute

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Variable analysis

independent variables

Introducing each of the 5002 shRNAs (targeting 957 genes)

dependent variables

The effect of introducing each of the 5002 shRNAs (targeting 957 genes) was determined in 19 cell lines
Expression profiling was used to generate a signature that correlates with KRAS activation and implicated NF-κB signaling in cell lines and tumors dependent on KRAS

control variables

The effect of introducing each of the 5002 shRNAs (targeting 957 genes) was normalized using the B-score metric
Secondary screen viability data was normalized using a percent of control statistic, given the biased nature of the candidate shRNA plate

positive controls

Not explicitly mentioned

negative controls

Not explicitly mentioned

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