The expression of Pap pili that facilitate the attach- ment of Escherichia coli to uroepithelial cells is shut off outside the host at temperatures below 268C. Ribo- nuclease protection analysis showed that this thermo- regulatory response was rapid as evidenced by the absence of papBA transcripts, coding for Pap pilin, after only one generation of growth at 238C. The his- tone-like nucleoid structuring protein H-NS and DNA sequences within papB were required for thermoregu- lation, but the PapB and PapI regulatory proteins were not. In vivo analysis of pap DNA methylation patterns indicated that H-NS or a factor regulated by H-NS bound within the pap regulatory region at 238C but not at 378C, as evidenced by H-NS-dependent inhibi- tion of methylation of the pap GATC sites designated GATC-I and GATC-II. These GATC sites lie upstream of the papBAp promoter and have been shown pre- viously to play a role in controlling Pap pili expression by regulating the binding of Lrp, a global regulator that is essential for activating papBAp transcription. Competitive electrophoretic mobility shift analysis showed that H-NS bound specifically to a pap DNA fragment containing the GATC-I and GATC-II sites. Moreover, H-NS blocked methylation of these pap GATC sites in vitro : H-NS blocked pap GATC methyla- tion at 1.4 mM but was unable to do so at higher con- centrations at which non-specific binding occurred. Thus, non-specific binding of H-NS to pap DNA was not sufficient to inhibit methylation of the pap GATC sites. These results suggest that the ability of H-NS to act as a methylation blocking factor is dependent upon the formation of a specific complex of H-NS with pap regulatory DNA. We hypothesize that a func- tion of H-NS such as oligomerization was altered at 238C, which enabled H-NS to repress pap gene expres- sion through the formation of a specific nucleoprotein complex.
White-Ziegler, Christine A.; Angus Hill, Melinda L.; Braaten, Bruce A.; Van Der Woude, Marjan W.; and Low, David A., "Thermoregulation of Escherichia coil pap Transcription: H-NS is a Temperature-Dependent DNA Methylation Blocking Factor" (1998). Biological Sciences: Faculty Publications, Smith College, Northampton, MA.