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. Author manuscript; available in PMC: 2014 Nov 8.
Published in final edited form as: Nat Commun. 2014 May 8;5:3795. doi: 10.1038/ncomms4795

Figure 2. S100A11 improves repair of large plasma membrane injuries.

Figure 2

(a) Quantification of MCF7-vector and -p95ErbB2 cells that failed to heal (TRITC-positive), 5 minutes following glass bead injury in the presence of FITC-dextran (labels all injured cells). When indicated, cells used were transfected for 72 h with indicated siRNAs. Error bars, SD for a triplicate representative (n = 3) experiment with 50-100 randomly chosen wounded cells / condition analyzed.

(b) Kinetics of cell membrane repair following laser injury. MCF7-vector and -p95ErbB2 cells incubated with FM1-43 dye were injured similarly by laser and the entry of FM1-43 was monitored by measuring FM1-43 intensity following injury (left). Error bars, SD for 10 independent cells / condition. Right panel shows representative images of cells before and after localized laser injury (white arrows). Note that unlike MCF7-vector cell, upon injury in MCF7-p95ErbB2 cell the damaged plasma membrane is excised and repaired at a new site marked by blue arrow.

(c) Images showing HeLa cells expressing S100A11-RFP and ANXA1-GFP (cell 1) or only ANXA1-GFP (cell 2) before and after localized laser injury (white arrows). Unlike cell 2 that does not overexpress S100A11, cell 1 repairs efficiently as indicated by the rapid clearance of ANXA1-GFP from the injury site; also see Video S2. Scale bar, 20 μm.

(d) FACS analysis of 7-AAD entry into uninjured and scrape injured HeLa cells allowed to repair for 10 minutes. Values indicate the percentage of cells in the gated area. The results are representative of three independent experiments.

(e) Viability of HeLa cells was analyzed by MTT reduction assay 96 h after transfection with control or S100A11 siRNAs (top left). Error bars, SD for three independent experiments.

Representative immunoblots of S100A11 (13 kDa) and Hsc70 (70 kDa; loading control) in lysates of HeLa cells transfected with indicated siRNAs 72 h prior to the lysis (bottom left). Representative phase contrast images of cells 96 h after transfection with indicated siRNAs (right). Scale bar 1mm. The asterisks represent P Values based on Students t-test: *** P < 0,001 when compared as indicated (A and B) or to the control siRNA-treated cells (E).