Fig. 6: βII-spectrin variants show different dynamic properties.

a Schematic representation of the βII-spectrin deletion variants analyzed in this study. In light green is shown the GFP tag, in dark green the actin-binding domain (ABD), and in red the PE/Ankyrin-binding site on spectrin repeats 14 and 15. Sequence alignment of PE/ANKbs between βI and βII-spectrin is shown: the mapped binding sites (red lines) are highly conserved between the two proteins. b Total cell lysates of MEFs expressing exogenous GFP-βII-spectrin variants analyzed by anti-GFP and anti-βII-spectrin antibodies in western blot assay (n = 2). c FRAP analysis of βII-spectrin deletion variants expressed in MEFs. In the graph are reported the normalized recovery curves (PE/ANKbs in the inset), while mobile fractions and half-time recoveries are plotted in the two graphs (d–e) (statistical analysis: one-way Anova with multiple comparison, *p = 0.0163, **p = 0.0071, ***p < 0.005, ****p < 0.0001, data are presented as mean values ± SD, n = 22 (FL), 13 (ΔABD), 15 (ΔPE/ANKbs), and 16 (PE/ANKbs) cells). f Normalized cell area growth during P2: three stereotypical MEFs transfected with GFP-βII-spectrin FL are plotted in blue, while MEFs expressing ∆ABD are shown in green and followed for 10 min after P1/P2 transition (T) by live TIRFM. g Quantification of ∆Area time⁻¹ extracted from subsequent frames in time lapses during 10 min after transition into P2 (FL n = 792 frames, ΔABD n = 840 frames, ΔPE/ANKbs n = 502 frames, n = 7–9 independent cells. Data are presented as mean values ± SD, statistical analysis one-way Anova with multiple comparison, **p = 0.0031, ****p < 0.0001).