title: The N-terminal amphipathic helices determine regulatory and effector functions of phage shock protein A (PspA) in Escherichia coli.
abstract: The phage shock protein (Psp) systems found in bacteria, archaea and higher plants respond to extracytoplasmic stresses that damage the cytoplasmic membrane and enable cells to repair their membranes. The conserved membrane-associated effector protein PspA has four α-helical domains (HD1-HD4) and helps to repair the membrane as a high-order oligomer. In enterobacteria, under non-stress conditions, PspA as a low-order assembly directly inhibits its cognate transcription activator PspF. Here we show that N-terminal amphipathic helices ahA and ahB in PspA HD1 are functional determinants involved in negative gene control and stress signal perception and its transduction via interactions with the PspBC membrane stress sensors and the inner membrane (IM). The amphipathic helices enable PspA to switch from a low-order gene regulator into an IM-bound high-order effector complex under membrane stress. Conserved residue proline 25 is involved in sequential use of the amphipathic helices and ahA-IM interaction. Single molecule imaging of eGFP-PspA and its amphipathic helices variants in live Escherichia coli cells show distinct spatial and temporal organisations of PspA corresponding to its negative control and effector functions. These findings inform studies on the role of the Psp system in persister cell formation and cell envelope protection in bacterial pathogens and provide a basis for exploring the specialised roles of PspA homologues such as YjfJ, LiaH and Vipp1.
authors.affiliation: Department of Life Sciences, Imperial College London, London SW7 2AZ, United Kingdom. Electronic address: g.jovanovic@imperial.ac.uk.
authors.forename: Goran
authors.initials: G
authors.lastname: Jovanovic
authors.affiliation: Department of Life Sciences, Imperial College London, London SW7 2AZ, United Kingdom.
authors.forename: Parul
authors.initials: P
authors.lastname: Mehta
authors.affiliation: Department of Life Sciences, Imperial College London, London SW7 2AZ, United Kingdom.
authors.forename: Christopher
authors.initials: C
authors.lastname: McDonald
authors.affiliation: Department of Life Sciences, Imperial College London, London SW7 2AZ, United Kingdom.
authors.forename: Anthony C
authors.initials: AC
authors.lastname: Davidson
authors.affiliation: Department of Life Sciences, Imperial College London, London SW7 2AZ, United Kingdom.
authors.forename: Povilas
authors.initials: P
authors.lastname: Uzdavinys
authors.affiliation: National Heart and Lung Institute, Imperial College London, London SW7 2AZ, United Kingdom.
authors.forename: Liming
authors.initials: L
authors.lastname: Ying
authors.affiliation: Department of Life Sciences, Imperial College London, London SW7 2AZ, United Kingdom. Electronic address: m.buck@imperial.ac.uk.
authors.forename: Martin
authors.initials: M
authors.lastname: Buck
mesh_terms: D001426:Bacterial Proteins / Q000737:chemistry* / Q000378:metabolism*; D004926:Escherichia coli / Q000378:metabolism*; D029968:Escherichia coli Proteins / Q000737:chemistry* / Q000378:metabolism*; D015964:Gene Expression Regulation, Bacterial; D006360:Heat-Shock Proteins / Q000737:chemistry* / Q000378:metabolism*; D017433:Protein Structure, Secondary
chemical_list: D001426:Bacterial Proteins; D029968:Escherichia coli Proteins; D006360:Heat-Shock Proteins; C069816:phage shock protein, Bacteria
keywords: PspA HD1; high-order oligomers; membrane stress; negative regulation of AAA(+); proline 25