Mechanistic insights into c-di-GMP-dependent control of the biofilm regulator FleQ from Pseudomonas aeruginosa

Author(s)

Matsuyama, Bruno Y. ; Krasteva, Petya V. ; Baraquet, Claudine ; Harwood, Caroline S. ; Sondermann, Holger ; Navarro, Marcos V. A. S.

• Document type: Journal article
• Source: PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA; v. 113, n. 2, p. 10-pg., 2016-01-12.

Abstract

Bacterial biofilm formation during chronic infections confers increased fitness, antibiotic tolerance, and cytotoxicity. In many pathogens, the transition from a planktonic lifestyle to collaborative, sessile biofilms represents a regulated process orchestrated by the intracellular second-messenger c-di-GMP. A main effector for c-di-GMP signaling in the opportunistic pathogen Pseudomonas aeruginosa is the transcription regulator FleQ. FleQ is a bacterial enhancer-binding protein (bEBP) with a central AAA+ ATPase sigma(54)-interaction domain, flanked by a C-terminal helix-turn-helix DNA-binding motif and a divergent N-terminal receiver domain. Together with a second ATPase, FleN, FleQ regulates the expression of flagellar and exopolysaccharide biosynthesis genes in response to cellular c-di-GMP. Here we report structural and functional data that reveal an unexpected mode of c-di-GMP recognition that is associated with major conformational rearrangements in FleQ. Crystal structures of FleQ's AAA+ ATPase domain in its apo-state or bound to ADP or ATP-gamma-S show conformations reminiscent of the activated ring-shaped assemblies of other bEBPs. As revealed by the structure of c-di-GMP-complexed FleQ, the second messenger interacts with the AAA+ ATPase domain at a site distinct from the ATP binding pocket. c-di-GMP interaction leads to active site obstruction, hexameric ring destabilization, and discrete quaternary structure transitions. Solution and cell-based studies confirm coupling of the ATPase active site and c-di-GMP binding, as well as the functional significance of crystallographic interprotomer interfaces. Taken together, our data offer unprecedented insight into conserved regulatory mechanisms of gene expression under direct c-di-GMP control via FleQ and FleQ-like bEBPs. (AU)

• FAPESP's process: 11/24168-2 - Structural and functional characterization of FleQ from Pseudomonas and Xanthomonas: a main transcriptional factor responsible for flagellar gene expression and biofilm formation
• Grantee: Bruno Yasui Matsuyama
• Support Opportunities: Scholarships in Brazil - Doctorate (Direct)
• FAPESP's process: 09/13238-0 - Structural and functional studies of proteins involved in c-di-GMP-mediated signalling pathways
• Grantee: Marcos Vicente de Albuquerque Salles Navarro
• Support Opportunities: Research Grants - Young Investigators Grants