Signal integration and transmission by the Cpx-TCS of Gram-negative bacteria

• Applicant: Professorin Dr. Sabine Hunke
• Subject Area: Metabolism, Biochemistry and Genetics of Microorganisms
• Term: from 2011 to 2016
• Project identifier: Deutsche Forschungsgemeinschaft (DFG) - Project number 204484694

Bacteria are equipped with two-component systems (TCS) to cope with environmental changes. However, the mechanistic details in TCS signal transduction are still poorly understood. The Cpx-TCS of Gram-negative bacteria is one of the best studied TCS and is composed of the sensor kinase CpxA, the response regulator CpxR, and the auxiliary protein CpxP. It responds to envelope stress which is induced by various signals e.g. pH, osmolarity or misfolded proteins. We have established that all three catalytic activities of the reconstituted sensor kinase CpxA can be modulated specifically by different signals: (i) CpxP inhibits autophosphorylation, (ii) a folding-deficient variant of periplasmic maltose binding protein stimulates transphosphorylation to CpxR, and (iii) lipids modulate dephosphorylation of phosphorylated CpxR. Moreover, our structural and functional studies on CpxP provide first insight how CpxP interacts with CpxA and serves as sensor for misfolded pilus subunits. I propose to continue our investigations on signal integration by the Cpx-TCS with special emphasis on the functions of CpxP: (i) inhibition of CpxA, (ii) pilus recognition and (iii) functional interaction with the periplasmic ATP-independent protease DegP. In addition and by taking advantage of our existing knowledge on signal integration by the Cpx-TCS, we will study one of the key questions in biology: How does a signal transverse a membrane? To this end, we have developed ‘Membrane-SPINE’, a method that will allow us to verify protein-protein interactions and have created a structural model of the catalytic core of CpxA in complex with CpxR. We will address critical residues for interaction and determine their relative reorientation during signalling by homobifunctional cross-linking and EPR after specific modulation of the catalytic activities of CpxA. Both subjects will contribute to the general understanding of signal integration and signal transmission.

• DFG Programme: Research Grants