AUTHOR=Panjaitan Novaria Sari Dewi , Horng Yu-Tze , Cheng Shih-Wen , Chung Wen-Ting , Soo Po-Chi 

TITLE=EtcABC, a Putative EII Complex, Regulates Type 3 Fimbriae via CRP-cAMP Signaling in Klebsiella pneumoniae

JOURNAL=Frontiers in Microbiology

VOLUME=10

YEAR=2019

URL=https://www.frontiersin.org/journals/microbiology/articles/10.3389/fmicb.2019.01558

DOI=10.3389/fmicb.2019.01558

ISSN=1664-302X

ABSTRACT=<p>Biofilm formation by <italic>Klebsiella pneumoniae</italic> on indwelling medical devices increases the risk of infection. Both type 1 and type 3 fimbriae are important factors in biofilm formation by <italic>K. pneumoniae</italic>. We found that a putative enzyme II (EII) complex of the phosphoenolpyruvate (PEP):carbohydrate phosphotransferase system (PTS), <italic>etcA</italic> (EIIA)-<italic>etcB</italic> (EIIB)-<italic>etcC</italic> (EIIC), regulated biofilm and type 3 fimbriae formation by <italic>K. pneumoniae</italic> STU1. In this study, the regulatory mechanism of <italic>etcABC</italic> in <italic>K. pneumoniae</italic> type 3 fimbriae formation was investigated. We found via quantitative RT-PCR that overexpression of <italic>etcABC</italic> enhanced the transcription level of the <italic>mrk</italic> operon, which is involved in type 3 fimbriae synthesis, and reduced the transcription level of the <italic>fim</italic> operon, which is involved in type 1 fimbriae synthesis. To gain further insight into the role of <italic>etcABC</italic> in type 3 fimbriae synthesis, we analyzed the region upstream of the <italic>mrk</italic> operon and found the potential cyclic 3′5′-adenosine monophosphate (cAMP) receptor protein (CRP) binding site. After <italic>crp</italic> was deleted in <italic>K. pneumoniae</italic> STU1 and two clinical isolates, these three <italic>crp</italic> mutant strains could not express MrkA, the major subunit of the fimbrial shaft, indicating that CRP positively regulated type 3 fimbriae synthesis. Moreover, a <italic>crp</italic> mutant overexpressing <italic>etcABC</italic> could not express MrkA, indicating that the regulation of type 3 fimbriae by <italic>etcABC</italic> was dependent on CRP. In addition, deletion of <italic>cyaA</italic>, which encodes the adenylyl cyclase that synthesizes cAMP, and deletion of <italic>crr</italic>, which encodes the glucose-specific EIIA, led to a reduction in <italic>lac</italic> operon regulation and therefore bacterial lactose uptake in <italic>K. pneumoniae</italic>. Exogenous cAMP but not <italic>etcABC</italic> overexpression compensated for the role of <italic>cyaA</italic> in bacterial lactose uptake. However, either <italic>etcABC</italic> overexpression or exogenous cAMP compensated for the role of <italic>crr</italic> in bacterial <italic>lac</italic> operon regulation that would eventually restore lactose uptake. We also found via ELISA and the <italic>luxCDABE</italic> reporter system that overexpression of <italic>etcABC</italic> increased intracellular cAMP levels and the transcription level of <italic>crp</italic>, respectively, in <italic>K. pneumoniae</italic>. In conclusion, overexpression of <italic>etcABC</italic> positively regulated cAMP production and cAMP-CRP activity to activate the <italic>mrk</italic> operon, resulting in increased type 3 fimbriae synthesis in <italic>K. pneumoniae</italic>.</p>