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Investigating the role of protein O‐fucosyltransferase 1 in Toll‐like receptor signaling (1004.6)
Author(s) -
Mann Jessica,
Li Ning,
Laky Karen,
Fraser Iain,
NitaLazar Aleksandra
Publication year - 2014
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.28.1_supplement.1004.6
Subject(s) - microbiology and biotechnology , signal transduction , biology , wnt signaling pathway , notch signaling pathway
Protein O‐fucosyltransferase 1 (Pofut1) adds fucose to the serine or threonine residues in the consensus sequence C2‐X(4,5)‐[S/T]‐C3 of EGF domains, where C2 and C3 are the second and third conserved cysteines. The most widely studied Pofut1 substrate is Notch, and the modification is essential in many developmental events involving Notch signaling, such as the development of the nervous system, myelopoiesis and lymphopoiesis, and later T‐cell and B‐cell development. It has not been, however, previously implicated in innate immune signaling. We have identified protein O‐fucosyltransferase 1 (Pofut1) as a significant hit in a genome‐wide RNAi screen of Toll‐like receptor (TLR) signaling in mouse RAW264.7 cells, where Pofut1 knockdown led to a reduction in LPS‐induced TNF expression. To further investigate the role of Pofut1 in the TLR signaling, we are using both Pofut1 siRNA transfected RAW264.7 cells and bone marrow derived macrophages (BMDM) from Pofut1 conditional knockout mice. We have studied the TLR4 pathway upon stimulation with lipopolysaccharide, combining cytokine expression assays, protein phosphorylation and protein expression profiling using Western blotting and mass spectrometry, and protein localization experiments. Grant Funding Source : This research was supported by the Intramural Research program of NIAID, NIH.