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. 2014 Jun 9;9(6):e98900. doi: 10.1371/journal.pone.0098900

Figure 8. Cysteines in the C-terminal region of syntaxin 11 are required for membrane association.

Figure 8

(A) C279, C280, C282, C283 and C285 (shown in red) were predicted by the CSS-PALM 2.04 software [38] to be potential S-acylation sites in syntaxin 11. The corresponding 5 cysteine residues were mutated to alanine to generate the de novo mutant syntaxin 11C5A. (B) Analysis of the membrane association of syntaxin 11C5A in YTS NK cells. Postnuclear supernatants from YTS NK cells were fractionated by centrifugation into pellet (membrane) and supernatant (cytosolic) fractions. The fractions were resolved by SDS-PAGE and GFP-syntaxin 11C5A was detected by probing immunoblots with a syntaxin 11 specific antibody. (C) Analysis of the localization of the syntaxin 11C5A mutant in YTS NK cells. YTS cells expressing GFP-syntaxin 11C5A were then imaged in the absence of target cells (Resting) or conjugated to 721.221 cells pre-stained with Cell Trace (blue in the merge image panels). Cells were imaged using a Zeiss LSM700 laser scanning confocal microscope. Scale bars 5 µm.