Screen for genes involved in radiation survival of Escherichia coli and construction of a reference database

摘要

A set of 3907 single-gene knockout (Keio collection) strains of Escherichia coli K-12 was examined for strains with increased susceptibility to killing by X- or UV-radiation. After screening with a high-throughput resazurin-based assay and determining radiation survival with triplicate clonogenic assays, we identified 76 strains (and associated deleted genes) showing statistically-significant increased radiation sensitivity compared to a control strain. To determine gene novelty, we constructed a reference database comprised of genes found in nine similar studies including ours. This database contains 455 genes comprised of 103 common genes (found 2-7 times), and 352 uncommon genes (found once). Our 76 genes includes 43 common genes and 33 uncommon (potentially novel) genes, i.e., appY, atoS, betB, bglj, clpP, cpxA, cysB, cysE, ddA, dgkA, dppF, dusB, elfG, eutK, fadD,glnA, grot, guaB, intF, prpR, queA, rplY, seqA, sufCyadC, yagj, yahD, yahO, ybaK, ybfA, yfaL, yhjV, and yiaL Of our 33 uncommon gene mutants, 4 (12%) were sensitive only to UV-radiation, 10 (30%) only to X-radiation, and 19 (58%) to both radiations. Our uncommon mutants vs. our common mutants showed more radiation specificity, i.e., 12% vs. 9% (sensitive only to UV-); 30% vs. 16% (X-) and 58% vs. 74% (both radiations). Considering just our radiation-sensitive mutants, the median UV-radiation survival (75 J m~2) for 23 uncommon mutants was 6.84E-3 compared to 1.85E-3 for 36 common mutants (P=0.025). Similarly, the average X-radiation survival for 29 uncommon mutants was 1.08E-2, compared to 6.19E-3 for 39 common mutants (P=0.010). Comparing gene functions using MultiFun terms, uncommon genes tended to show less involvement in DNA repair-relevant categories (information transfer and cell processes), but greater involvement in seven other categories. Our analysis of 455 genes suggests cell survival and DNA repair processes are more complex than previously understood, and may be compromised by deficiencies in other processes.