Functions that protect Escherichia coli from DNA-protein crosslinks.

Journal Article (Journal Article)

Pathways for tolerating and repairing DNA-protein crosslinks (DPCs) are poorly defined. We used transposon mutagenesis and candidate gene approaches to identify DPC-hypersensitive Escherichia coli mutants. DPCs were induced by azacytidine (aza-C) treatment in cells overexpressing cytosine methyltransferase; hypersensitivity was verified to depend on methyltransferase expression. We isolated hypersensitive mutants that were uncovered in previous studies (recA, recBC, recG, and uvrD), hypersensitive mutants that apparently activate phage Mu Gam expression, and novel hypersensitive mutants in genes involved in DNA metabolism, cell division, and tRNA modification (dinG, ftsK, xerD, dnaJ, hflC, miaA, mnmE, mnmG, and ssrA). Inactivation of SbcCD, which can cleave DNA at protein-DNA complexes, did not cause hypersensitivity. We previously showed that tmRNA pathway defects cause aza-C hypersensitivity, implying that DPCs block coupled transcription/translation complexes. Here, we show that mutants in tRNA modification functions miaA, mnmE and mnmG cause defects in aza-C-induced tmRNA tagging, explaining their hypersensitivity. In order for tmRNA to access a stalled ribosome, the mRNA must be cleaved or released from RNA polymerase. Mutational inactivation of functions involved in mRNA processing and RNA polymerase elongation/release (RNase II, RNaseD, RNase PH, RNase LS, Rep, HepA, GreA, GreB) did not cause aza-C hypersensitivity; the mechanism of tmRNA access remains unclear.

Full Text

Duke Authors

Cited Authors

  • Krasich, R; Wu, SY; Kuo, HK; Kreuzer, KN

Published Date

  • April 2015

Published In

Volume / Issue

  • 28 /

Start / End Page

  • 48 - 59

PubMed ID

  • 25731940

Pubmed Central ID

  • PMC4385401

Electronic International Standard Serial Number (EISSN)

  • 1568-7856

Digital Object Identifier (DOI)

  • 10.1016/j.dnarep.2015.01.016


  • eng

Conference Location

  • Netherlands