Fig. 5. Phosphatase-inhibitor-induced keratin filament disruption in differentiated keratinocytes proceeds faster in the absence of epiplakin. (A) Primary wild-type (+/+) and epiplakin-deficient (–/–) keratinocytes differentiated in medium containing 1.3 mM Ca²⁺ were treated with OA for 2, 4 and 6 hours and immunolabeled with antibodies to pan-keratin. Arrows indicate examples of cells with partially disrupted keratin filament networks. Arrowheads indicate examples of cells with totally disrupted keratin networks. Scale bar: 10 µm. (B) Decrease of cells with intact filaments during the first 2 hours after onset of OA-induced filament breakdown (left panel) and increase of cells with totally disrupted filament networks over a 2 hour period, starting when the first cells of this type appeared (right panel). Mean values ± s.d. of four independent experiments are based on >100 cells evaluated per time point. (C) Statistical analysis of keratinocytes with complete, partial or no filament breakdown upon okadaic acid incubation. Note that the trend of faster keratin network collapse in epiplakin^–/– compared with wild type (+/+) cells was observed in four independent experimental series, regardless of the genetic background of the mice used as cell donors. One representative series is shown. Relative proportions of intact (filamentous), partially disrupted, and disrupted keratin filament networks are based on >100 cells evaluated per time point. (D) Increase of cells without intact filament networks upon incubation with orthovanadate over a 9 minute period. Mean values ± s.d. of cells without intact filaments per time-point are based on >100 cells evaluated per time point.