Figure 1.

The stability of lamin B1 interactions is dependent on posttranslational modification by Rce1. (a) FLIP in WT mouse embryonic fibroblasts, Rce^−/−, and Icmt^−/− cells expressing GFP–lamin B1. A specific ROI was photobleached at full laser power for 250 s. The postbleach image shows the extent of fluorescence loss in the nonbleached area, which is indicative of the stability of GFP–lamin B1 at the nuclear periphery. Bar, 10 μm. (b) Quantitative FLIP. An ROI outside the photobleached area shown in panel a was used to measure fluorescence loss after photobleaching (mean values ± SD; n = 5). The background intensity was subtracted and the values normalized by dividing by the intensities of an ROI in an adjacent control cell to account for photobleaching of the 5% laser power used for scanning the nonbleached areas. GFP–lamin B1 interactions at the nuclear periphery are more stable in WT and Icmt^−/− (c) than in Rce^−/− (b), indicating that carboxymethylation has little or no effect on the stability of the nuclear lamina. (d) FLIP of GFP–lamin B1 in the Lmnb1^−/− cells used in the current study does not show any significant difference from its behavior in WT cells, indicating that the difference observed in the Rce1^−/− cells is due to a lack of processing and not another cell-specific factor.