A Putative Bacterial ABC Transporter Circumvents the Essentiality of Signal Peptidase.
The type I signal peptidase of Staphylococcus aureus, SpsB, is an attractive antibacterial target because it is essential for viability and extracellularly accessible. We synthesized compound 103, a novel arylomycin-derived inhibitor of SpsB with significant potency against various clinical S. aureus strains (MIC of ~1 µg/ml). The predominant clinical strain USA300 developed spontaneous resistance to compound 103 with high frequency, resulting from single point mutations inside or immediately upstream of cro/cI, a homolog of the lambda phage transcriptional repressor cro These cro/cI mutations led to marked (>50-fold) overexpression of three genes encoding a putative ABC transporter. Overexpression of this ABC transporter was both necessary and sufficient for resistance and, notably, circumvented the essentiality of SpsB during in vitro culture. Mutation of its predicted ATPase gene abolished resistance, suggesting a possible role for active transport; in these bacteria, resistance to compound 103 occurred with low frequency and through mutations in spsB Bacteria overexpressing the ABC transporter and lacking SpsB were capable of secreting a subset of proteins that are normally cleaved by SpsB and instead were cleaved at a site distinct from the canonical signal peptide. These bacteria secreted reduced levels of virulence-associated proteins and were unable to establish infection in mice. This study reveals the mechanism of resistance to a novel arylomycin derivative and demonstrates that the nominal essentiality of the S. aureus signal peptidase can be circumvented by the upregulation of a putative ABC transporter in vitro but not in vivo



The type I signal peptidase of Staphylococcus aureus (SpsB) enables the secretion of numerous proteins by cleavage of the signal peptide. We synthesized an SpsB inhibitor with potent activity against various clinical S. aureus strains. The predominant S. aureus strain USA300 develops resistance to this inhibitor by mutations in a novel transcriptional repressor (cro/cI), causing overexpression of a putative ABC transporter. This mechanism promotes the cleavage and secretion of various proteins independently of SpsB and compensates for the requirement of SpsB for viability in vitro However, bacteria overexpressing the ABC transporter and lacking SpsB secrete reduced levels of virulence-associated proteins and are unable to infect mice. This study describes a bacterial resistance mechanism that provides novel insights into the biology of bacterial secretion.
Department of Infectious Diseases, Genentech, Inc., South San Francisco, California, USA.
J Hiroshi
JH
Morisaki
Department of Infectious Diseases, Genentech, Inc., South San Francisco, California, USA.
Peter A
PA
Smith
Department of Infectious Diseases, Genentech, Inc., South San Francisco, California, USA.
Shailesh V
SV
Date
Department of Infectious Diseases, Genentech, Inc., South San Francisco, California, USA.
Kimberly K
KK
Kajihara
Department of Infectious Diseases, Genentech, Inc., South San Francisco, California, USA.
Chau Linda
CL
Truong
Department of Molecular Biology, Genentech, Inc., South San Francisco, California, USA.
Zora
Z
Modrusan
Department of Translational Immunology, Genentech, Inc., South San Francisco, California, USA.
Donghong
D
Yan
Department of Translational Immunology, Genentech, Inc., South San Francisco, California, USA.
Jing
J
Kang
Department of Translational Immunology, Genentech, Inc., South San Francisco, California, USA.
Min
M
Xu
Department of Infectious Diseases, Genentech, Inc., South San Francisco, California, USA.
Ishita M
IM
Shah
Department of Biochemical and Cellular Pharmacology, Genentech, Inc., South San Francisco, California, USA.
Robert
R
Mintzer
Department of Infectious Diseases, Genentech, Inc., South San Francisco, California, USA.
Eric M
EM
Kofoed
Department of Protein Chemistry, Genentech, Inc., South San Francisco, California, USA.
Tommy K
TK
Cheung
Department of Protein Chemistry, Genentech, Inc., South San Francisco, California, USA.
David
D
Arnott
Department of Discovery Chemistry, Genentech, Inc., South San Francisco, California, USA.
Michael F T
MF
Koehler
Department of Infectious Diseases, Genentech, Inc., South San Francisco, California, USA Department of Biochemical and Cellular Pharmacology, Genentech, Inc., South San Francisco, California, USA.
Christopher E
CE
Heise
Department of Infectious Diseases, Genentech, Inc., South San Francisco, California, USA.
Eric J
EJ
Brown
Department of Infectious Diseases, Genentech, Inc., South San Francisco, California, USA whazenbos@yahoo.com tan.man-wah@gene.com.
Man-Wah
MW
Tan
Department of Infectious Diseases, Genentech, Inc., South San Francisco, California, USA whazenbos@yahoo.com tan.man-wah@gene.com.
Wouter L W
WL
Hazenbos
D018528:ATP-Binding Cassette Transporters / Q000235:genetics* / Q000378:metabolism*; D000818:Animals; D000900:Anti-Bacterial Agents / Q000494:pharmacology; D004195:Disease Models, Animal; D024881:Drug Resistance, Bacterial; D015870:Gene Expression; D008565:Membrane Proteins / Q000037:antagonists & inhibitors / Q000235:genetics* / Q000378:metabolism*; D051379:Mice; D008826:Microbial Sensitivity Tests; D009154:Mutation; D012641:Selection, Genetic; D012697:Serine Endopeptidases / Q000235:genetics* / Q000378:metabolism*; D013203:Staphylococcal Infections / Q000382:microbiology; D013211:Staphylococcus aureus / Q000187:drug effects / Q000201:enzymology* / Q000235:genetics* / Q000378:metabolism; D014774:Virulence
D018528:ATP-Binding Cassette Transporters; D000900:Anti-Bacterial Agents; D008565:Membrane Proteins; D012697:Serine Endopeptidases; C026515:type I signal peptidase