Figure 5. Dye transfer assays for gap junction forming mutants.Intercellular communication was assayed by scrape dye loading using Lucifer Yellow (LY) and Dextran Texas Red (TxR). LY will pass through gap junctions while Dextran TxR will not. (Panel A) Confluent gap-junction deficient parental HeLa cells (left column) and HeLa cells transiently transfected with Cx26 WT-GFP-4C (right column) serve as negative and positive controls, respectively. Top row: Images of LY fluorescence. Middle row: Images of TxR fluorescence Bottom row: Differential Interference Contrast (DIC) light micrograph of the same field of cells showing cell-cell contact. Dextran Texas Red acts as a reporter for dead or non-communicating cells and those along the scratch that also contained Lucifer Yellow were not counted in this analysis. (Panel B) Similar images as in (Panel A) for the two mutants, M195T and A197S, which made detergent stable hemichannels and channels. (Panel C) LY (top), TxR (middle) and DIC images (bottom) for mutants that made detergent unstable hemichannels and channels (left to right, C202F, I203T, L205V and N206S). The inset at the left is a 4.5x magnification of a LY non-transferring cell expressing C202F gap junctions. The arrow points to these cells in the C202F LY image. (D) Histogram of levels of communicating cells for each mutant. Error bars are standard errors of the mean (SEM).
Image published in: Ambrosi C et al. (2013)
Image reproduced on Xenbase with permission of the publisher and the copyright holder. Creative Commons Attribution license
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