The small regulatory proteins of the PhoQ/PhoP two-component system

• Applicant: Dr. Jing Yuan
• Subject Area: Metabolism, Biochemistry and Genetics of Microorganisms ; Medical Microbiology and Mycology, Hygiene, Molecular Infection Biology
• Term: since 2020
• Project identifier: Deutsche Forschungsgemeinschaft (DFG) - Project number 453206916

Increasing number of small proteins have been discovered in the past a few years in all three domains of life. A small portion of them have known functions and participate in diverse aspects of cellular processes and bacterial communities. In Escherichia coli, about one third of small proteins are predicted as membrane proteins, suggesting their possible roles in sensing environmental changes and mediating cellular responses. The bacterial two-component systems are the primary signaling pathways that connect environmental input stimuli to cell physiological responses. The PhoQ/PhoP two-component system is essential for enterobacteria to sense host-associated stimuli, control bacterial virulence and initiate specific stress responses. Two membrane-localized small proteins, MgrB and SafA, directly regulate the activity of PhoQ sensor kinase and influence the fitness and pathogenicity of enterobacteria. Disruption of the mgrB gene was shown to strongly correlate with resistance to the last resort antibiotic colistin. The small protein MgrB inhibits PhoQ kinase activity. In contrast, SafA activates PhoQ. Recently, our detailed study of MgrB identified 11 functionally essential amino acid residues spread across all regions of the protein. We pinpointed that a tryptophan residue in the transmembrane helix is crucial for PhoQ/MgrB complex formation and propose that the short cytosolic region of MgrB affects signal transduction in PhoQ. However, the exact mechanism of MgrB inhibition remains elusive. Additionally, our studies on PhoQ/MgrB complex showed reduced interaction at low magnesium directly supporting the notion that PhoQ/MgrB interaction can be modulated by environmental conditions. This observation reveals a new aspect of regulation, which will be systematically investigated for both MgrB and SafA in this project. Here, we propose to elucidate the mechanisms of MgrB and the dynamics of how MgrB and SafA regulate PhoQ activity using site-specific crosslinking, FRET and other complementary methods in vivo and in vitro. we aim to (i) uncover the molecular details of MgrB/PhoQ interaction, (ii) identify and characterize environmental conditions that modulate PhoQ/small protein interactions, (iii) study how PhoQ integrates signals from MgrB and SafA, (iv) explore additional protein targets and functions of MgrB, and (v) develop new antimicrobial agents based on MgrB. Our project is expected to contribute to the better understanding of the complex regulatory network of two-component systems, shed light on the physiological importance of small proteins on bacterial virulence, and generate a set of antimicrobial agents, which could provide additional means to address the worldwide increasing cases of colistin resistance.

• DFG Programme: Priority Programmes
• Subproject of: SPP 2002: Small Proteins in Prokaryotes, an Unexplored World