Linked Life Data

21220426

Source:http://linkedlifedata.com/resource/pubmed/id/21220426

Statements in which the resource exists as a subject.
PredicateObject
rdf:type
lifeskim:mentions
pubmed:issue
13
pubmed:dateCreated
2011-4-1
pubmed:abstractText
In the TLR4 signaling pathways, we previously characterized a signal regulator, LRRFIP2, that modulates the time course-dependent changes in NF-?B activity through its dynamic interaction with the TLR adaptor protein, MyD88. However, little is known about the driving force behind the LPS-inducible dynamics between LRRFIP2 and MyD88. We have therefore designed a multiplex label-free quantitative proteomics method to investigate dynamic changes of LRRFIP2 phosphorylation upon LPS stimulation. Given our observation that LRRFIP2 binds to MyD88 through its serine-rich domain in which most of serine residues have the propensity to be phosphorylated, we used collision-activated dissociation- and electron transfer dissociation-based methods in a complementary manner to unambiguously localize phosphorylation sites in the peptides constituting the serine-rich domain. Among 23 phosphorylation sites identified and first quantified by the label-free approach and then verified by the AACT/SILAC (amino acid-coded tagging/stable isotope labeling in cell culture)-based quantitation method, phosphorylation at serine 202 showed a significant LPS-induced dynamic change during the full-course cellular response to LPS stimulation. The substitution of serine 202 with nonphosphorylated residues by site-directed mutagenesis resulted in a weakened LRRFIP2-MyD88 interaction and a concurrently reduced activity in downstream NF-?B. Taking these results together, phosphorylation at serine 202 was found to regulate the dynamics of the LRRFIP2-MyD88 interaction, which in turn modulated the strength and duration of TLR4 signaling. Strategically, we have demonstrated the importance of precise identification of the biologically relevant phosphorylation site(s) using comprehensive mass spectrometry-based quantitative proteomics approaches in guiding downstream biological characterization experiments, which could otherwise be both time- and cost-consuming for a large number of phosphorylation possibilities.
pubmed:grant
pubmed:language
eng
pubmed:journal
pubmed:citationSubset
IM
pubmed:chemical
http://linkedlifedata.com/resource/pubmed/chemical/Carrier Proteins, http://linkedlifedata.com/resource/pubmed/chemical/LRRFIP2 protein, human, http://linkedlifedata.com/resource/pubmed/chemical/Lipopolysaccharides, http://linkedlifedata.com/resource/pubmed/chemical/MYD88 protein, human, http://linkedlifedata.com/resource/pubmed/chemical/Myd88 protein, mouse, http://linkedlifedata.com/resource/pubmed/chemical/Myeloid Differentiation Factor 88, http://linkedlifedata.com/resource/pubmed/chemical/NF-kappa B, http://linkedlifedata.com/resource/pubmed/chemical/TLR4 protein, human, http://linkedlifedata.com/resource/pubmed/chemical/Tlr4 protein, mouse, http://linkedlifedata.com/resource/pubmed/chemical/Toll-Like Receptor 4, http://linkedlifedata.com/resource/pubmed/chemical/lipopolysaccharide, Escherichia...
pubmed:status
MEDLINE
pubmed:month
Apr
pubmed:issn
1083-351X
pubmed:author
pubmed:issnType
Electronic
pubmed:day
1
pubmed:volume
286
pubmed:owner
NLM
pubmed:authorsComplete
Y
pubmed:pagination
10897-910
pubmed:meshHeading
pubmed:year
2011
pubmed:articleTitle
Unambiguous characterization of site-specific phosphorylation of leucine-rich repeat Fli-I-interacting protein 2 (LRRFIP2) in Toll-like receptor 4 (TLR4)-mediated signaling.
pubmed:affiliation
Program in Molecular Biology and Biotechnology, University of North Carolina, Chapel Hill, North Carolina 27599, USA.
pubmed:publicationType
Journal Article, Research Support, N.I.H., Extramural