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Genome-Wide Expression Profiling and Mutagenesis Studies Reveal that Lipopolysaccharide Responsiveness Appears To Be Absolutely Dependent on TLR4 and MD-2 Expression and Is Dependent upon Intermolecular Ionic Interactions

Meng, Jianmin; Gong, Mei; Björkbacka, Harry LU and Golenbock, Douglas T. (2011) In Journal of Immunology 187(7). p.3683-3693
Abstract
Lipid A (a hexaacylated 1,4' bisphosphate) is a potent immune stimulant for TLR4/MD-2. Upon lipid A ligation, the TLR4/MD-2 complex dimerizes and initiates signal transduction. Historically, studies also suggested the existence of TLR4/MD-2-independent LPS signaling. In this article, we define the role of TLR4 and MD-2 in LPS signaling by using genome-wide expression profiling in TLR4- and MD-2-deficient macrophages after stimulation with peptidoglycan-free LPS and synthetic Escherichia coli lipid A. Of the 1396 genes significantly induced or repressed by any one of the treatments in the wild-type macrophages, none was present in the TLR4-or MD-2-deficient macrophages, confirming that the TLR4/MD-2 complex is the only receptor for... (More)
Lipid A (a hexaacylated 1,4' bisphosphate) is a potent immune stimulant for TLR4/MD-2. Upon lipid A ligation, the TLR4/MD-2 complex dimerizes and initiates signal transduction. Historically, studies also suggested the existence of TLR4/MD-2-independent LPS signaling. In this article, we define the role of TLR4 and MD-2 in LPS signaling by using genome-wide expression profiling in TLR4- and MD-2-deficient macrophages after stimulation with peptidoglycan-free LPS and synthetic Escherichia coli lipid A. Of the 1396 genes significantly induced or repressed by any one of the treatments in the wild-type macrophages, none was present in the TLR4-or MD-2-deficient macrophages, confirming that the TLR4/MD-2 complex is the only receptor for endotoxin and that both are required for responses to LPS. Using a molecular genetics approach, we investigated the mechanism of TLR4/MD-2 activation by combining the known crystal structure of TLR4/MD-2 with computer modeling. According to our murine TLR4/MD-2-activation model, the two phosphates on lipid A were predicted to interact extensively with the two positively charged patches on mouse TLR4. When either positive patch was abolished by mutagenesis into Ala, the responses to LPS and lipid A were nearly abrogated. However, the MyD88-dependent and -independent pathways were impaired to the same extent, indicating that the adjuvant activity of monophosphorylated lipid A most likely arises from its decreased potential to induce an active receptor complex and not more downstream signaling events. Hence, we concluded that ionic interactions between lipid A and TLR4 are essential for optimal LPS receptor activation. The Journal of Immunology, 2011, 187: 3683-3693. (Less)
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author
organization
publishing date
type
Contribution to journal
publication status
published
subject
in
Journal of Immunology
volume
187
issue
7
pages
3683 - 3693
publisher
American Association of Immunologists
external identifiers
  • wos:000295036400027
  • scopus:80053487700
ISSN
1550-6606
DOI
10.4049/jimmunol.1101397
language
English
LU publication?
yes
id
20404883-d1b3-4b9d-82ff-3601c4dc98d6 (old id 2272024)
date added to LUP
2012-01-02 09:47:49
date last changed
2017-07-02 04:14:04
@article{20404883-d1b3-4b9d-82ff-3601c4dc98d6,
  abstract     = {Lipid A (a hexaacylated 1,4' bisphosphate) is a potent immune stimulant for TLR4/MD-2. Upon lipid A ligation, the TLR4/MD-2 complex dimerizes and initiates signal transduction. Historically, studies also suggested the existence of TLR4/MD-2-independent LPS signaling. In this article, we define the role of TLR4 and MD-2 in LPS signaling by using genome-wide expression profiling in TLR4- and MD-2-deficient macrophages after stimulation with peptidoglycan-free LPS and synthetic Escherichia coli lipid A. Of the 1396 genes significantly induced or repressed by any one of the treatments in the wild-type macrophages, none was present in the TLR4-or MD-2-deficient macrophages, confirming that the TLR4/MD-2 complex is the only receptor for endotoxin and that both are required for responses to LPS. Using a molecular genetics approach, we investigated the mechanism of TLR4/MD-2 activation by combining the known crystal structure of TLR4/MD-2 with computer modeling. According to our murine TLR4/MD-2-activation model, the two phosphates on lipid A were predicted to interact extensively with the two positively charged patches on mouse TLR4. When either positive patch was abolished by mutagenesis into Ala, the responses to LPS and lipid A were nearly abrogated. However, the MyD88-dependent and -independent pathways were impaired to the same extent, indicating that the adjuvant activity of monophosphorylated lipid A most likely arises from its decreased potential to induce an active receptor complex and not more downstream signaling events. Hence, we concluded that ionic interactions between lipid A and TLR4 are essential for optimal LPS receptor activation. The Journal of Immunology, 2011, 187: 3683-3693.},
  author       = {Meng, Jianmin and Gong, Mei and Björkbacka, Harry and Golenbock, Douglas T.},
  issn         = {1550-6606},
  language     = {eng},
  number       = {7},
  pages        = {3683--3693},
  publisher    = {American Association of Immunologists},
  series       = {Journal of Immunology},
  title        = {Genome-Wide Expression Profiling and Mutagenesis Studies Reveal that Lipopolysaccharide Responsiveness Appears To Be Absolutely Dependent on TLR4 and MD-2 Expression and Is Dependent upon Intermolecular Ionic Interactions},
  url          = {http://dx.doi.org/10.4049/jimmunol.1101397},
  volume       = {187},
  year         = {2011},
}