title: Patch clamp characterization of the effect of cardiolipin on MscS of E. coli.
abstract: The bacterial mechanosensitive channels MscS and MscL are gated by an increase in membrane tension when the bacterium experiences hypoosmotic shock. It has been well established that membrane lipids modulate the mechanosensitivity and gating behavior of these channels. The focus of this study is a negatively charged phospholipid, cardiolipin, which has been shown to localize at curved regions of the bacterial cell, including the poles and the septum, and to have a strong preference for binding to membrane proteins. Here we characterize the effect of cardiolipin on MscS, the mechanosensitive channel of small conductance, using patch-clamp electrophysiology. We compare the gating kinetics and mechanosensitivity of the channel in both azolectin and mixtures of pure lipids DOPE/DOPC liposomes with and without cardiolipin. In azolectin liposomes, the addition of 10 % cardiolipin abolishes hysteresis of MscS, but MscL remains largely unaffected, indicating that cardiolipin may stabilize the closed state of MscS. On the other hand, mixtures of DOPE/DOPC abolish the hysteresis gating of MscS even in the absence of cardiolipin, and the addition of cardiolipin increases the opening and closing thresholds of both MscS and MscL. In addition, we show that MscS gates more frequently when cardiolipin is present in both the azolectin and pure lipid systems; this dose-dependent effect ultimately destabilizes the open state of MscS and we consider the functional implications of this cardiolipin effect in the bacterial osmotic response. Our results show that cardiolipin modulates the mechanosensitivity and gating characteristics of MscS, indicating its important role in the physiology of bacterial cells.
authors.affiliation: Molecular Cardiology and Biophysics Division, Victor Chang Cardiac Research Institute, 405 Liverpool St, Darlinghurst, NSW, 2010, Australia.
authors.forename: Pietro
authors.initials: P
authors.lastname: Ridone
authors.forename: Yoshitaka
authors.initials: Y
authors.lastname: Nakayama
authors.forename: Boris
authors.initials: B
authors.lastname: Martinac
authors.forename: Andrew R
authors.initials: AR
authors.lastname: Battle
mesh_terms: D000200:Action Potentials / Q000187:drug effects; D000595:Amino Acid Sequence; D002308:Cardiolipins / Q000494:pharmacology*; D029968:Escherichia coli Proteins / Q000737:chemistry / Q000378:metabolism*; D015640:Ion Channel Gating / Q000187:drug effects*; D007473:Ion Channels / Q000737:chemistry / Q000378:metabolism*; D008081:Liposomes / Q000737:chemistry / Q000378:metabolism; D008969:Molecular Sequence Data; D018408:Patch-Clamp Techniques
chemical_list: D002308:Cardiolipins; D029968:Escherichia coli Proteins; D007473:Ion Channels; D008081:Liposomes; C086596:MscL protein, E coli; C469542:MscS protein, E coli
references.citation: Nat Struct Biol. 2002 Sep;9(9):696-703
references.pmid: 12172537
references.citation: Eur Biophys J. 2009 Apr;38(4):415-25
references.pmid: 19132368
references.citation: J Gen Physiol. 1999 Apr;113(4):525-40
references.pmid: 10102934
references.citation: J Bacteriol. 2010 Feb;192(4):912-24
references.pmid: 20008071
references.citation: Proc Natl Acad Sci U S A. 2014 Sep 23;111(38):13864-9
references.pmid: 25201991
references.citation: Biochim Biophys Acta. 2009 Oct;1788(10):2069-79
references.pmid: 19328771
references.citation: Biophys J. 1989 Sep;56(3):631-6
references.pmid: 2477074
references.citation: Proc Natl Acad Sci U S A. 1999 Aug 3;96(16):8890-4
references.pmid: 10430866
references.citation: Proc Natl Acad Sci U S A. 2003 Dec 23;100(26):15959-64
references.pmid: 14671322
references.citation: Cell Physiol Biochem. 2011;28(6):1051-60
references.pmid: 22178995
references.citation: EMBO J. 1999 Apr 1;18(7):1730-7
references.pmid: 10202137
references.citation: Eur Biophys J. 2005 Jul;34(5):434-41
references.pmid: 15834558
references.citation: Nat Cell Biol. 2003 Mar;5(3):175-8
references.pmid: 12646867
references.citation: J Bacteriol. 2012 Sep;194(18):4802-9
references.pmid: 22685280
references.citation: Biochemistry. 2009 Dec 1;48(47):11185-95
references.pmid: 19863062
references.citation: Biophys J. 2002 Jul;83(1):290-8
references.pmid: 12080120
references.citation: Nature. 2014 Jun 5;510(7503):172-175
references.pmid: 24899312
references.citation: Antioxid Redox Signal. 2014 Feb 20;20(6):952-69
references.pmid: 23834368
references.citation: J Gen Physiol. 2001 Aug;118(2):193-206
references.pmid: 11479346
references.citation: Biochemistry. 2007 Jun 12;46(23):6766-73
references.pmid: 17500538
references.citation: Biophys J. 2004 Apr;86(4):2113-20
references.pmid: 15041651
references.citation: Biophys J. 2010 Nov 3;99(9):2870-8
references.pmid: 21044584
references.citation: Proc Natl Acad Sci U S A. 2011 Apr 12;108(15):6264-9
references.pmid: 21444798
references.citation: J Bacteriol. 2014 Oct;196(19):3386-98
references.pmid: 25002539
references.citation: Science. 2008 Aug 29;321(5893):1210-4
references.pmid: 18755978
references.citation: Biochim Biophys Acta. 2009 Oct;1788(10):2092-100
references.pmid: 19539601
references.citation: J Bacteriol. 2005 Mar;187(5):1591-603
references.pmid: 15716429
references.citation: Antioxid Redox Signal. 2014 Apr 20;20(12):1925-53
references.pmid: 24094094
references.citation: J Cell Sci. 2014 Jan 1;127(Pt 1):11-9
references.pmid: 24345373
references.citation: Biophys J. 2007 Dec 1;93(11):3884-99
references.pmid: 17704167
references.citation: J Membr Biol. 1996 May;151(2):175-87
references.pmid: 8661505
references.citation: FEBS Lett. 2009 Jan 22;583(2):407-12
references.pmid: 19111548
references.citation: Proc Natl Acad Sci U S A. 2012 May 29;109(22):8770-5
references.pmid: 22586095
references.citation: J Biol Chem. 1995 Aug 4;270(31):18329-34
references.pmid: 7543101
references.citation: Chem Phys Lipids. 1999 Aug;101(1):45-56
references.pmid: 10810924
references.citation: J Biol Chem. 2008 May 2;283(18):12314-23
references.pmid: 18326496
references.citation: Biochemistry. 2013 Aug 13;52(32):5415-20
references.pmid: 23875651
references.citation: Channels (Austin). 2014;8(1):5-12
references.pmid: 24262975