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. 2020 Nov 19;8:607080. doi: 10.3389/fcell.2020.607080

FIGURE 12.

FIGURE 12

O-GlcNAcylation controls the interaction of ICln with α-integrin. (A) NIH/3T3 cells were transfected for 48 h with the FRET acceptor αIIbshort-EYFP and ECFP-ICln wild type or Ser67Ala, or with ECFP as a control, and imaged before (pre) and after (post) photobleaching of the FRET acceptor. FRET efficiency was determined in three–four regions of interest (ROIs) of the plasma membrane of each cell. (B) The O-GlcNAcylation levels were left unmodified or (C) increased by incubation of cells with 100 μM PUGNAc plus 5 mM glucosamine for 1 h. The vehicle was 0.1% DMSO plus 5 mM D-glucose. (D) The O-GlcNAcylation levels were reduced by co-transfection of cells with OGA-L and the transfection marker dsRed as separated proteins. (n) refers to the number of ROIs of cells from at least four subcultures. ***p < 0.001, **p < 0.01, *p < 0.05, one-way ANOVA with Dunnett’s post-test. FRET, fluorescence resonance energy transfer; EYFP, enhanced yellow fluorescent protein; ECFP, enhanced cyan fluorescent protein; DMSO, dimethyl sulfoxide.