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Fig. 5 | Breast Cancer Research

Fig. 5

From: Quantitative glycoproteomics analysis identifies novel FUT8 targets and signaling networks critical for breast cancer cell invasiveness

Fig. 5

FUT8 modulates core fucosylation of IL6ST protein and its downstream signaling. a and b IL6ST and OSM receptor (OSMR) proteins core fucosylated by FUT8 in HEK-293 T cells. Recombinant IL6ST or OSMR proteins in control or two FUT8-KO 293 T cell lines were probed with biotinylated LCA, then detected with streptavidin-conjugated horseradish peroxidase. Protein domain organization of these selected proteins is illustrated according to Uniprot data (http://uniprot.org). Amino acid number and potential N-linked glycosylation sites are marked on the upper and lower side, respectively. Core-fucosylated sites confirmed by LC–MS/MS analyses are in red. TM, transmembrane domain. c Core-fucosylation sites of IL6ST protein. Shows LC–MS/MS summed extracted ion chromatogram (XIC) of identified core-fucosylated glycopeptides for IL6ST. The number on asparagine (N) indicates the position in the protein sequence. d FUT8 KO impaired IL-6 and OSM signaling in STAT3 reporter HEK cells. Parental or FUT8-KO HEK-Blue™ IL-6 cells (InvivoGen) were treated with recombinant IL-6 (upper) or OSM (lower). Levels of STAT3-inducible secreted embryonic alkaline phosphatase (SEAP) indicating STAT3 activity were monitored by using QUANTI-Blue. Data are mean ± SD. **p < 0.01, *p < 0.05, FUT8-KO #1 or FUT8-KO#2 versus parental HEK-Blue™ IL-6 cells. e FUT8 KO impaired IL-6 and OSM signaling in MDA-MB-231 cells. Control and FUT8-KO MDA-MB-231 cells were treated with the indicated concentrations of recombinant IL-6 (left panel) or OSM (right panel) for 15 min. Cell lysates underwent western blot analysis with antibody for phosphorylated STAT3 or total STAT3

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