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. 2013 Mar 4;200(5):567–576. doi: 10.1083/jcb.201211068

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© 2013 Yi et al.

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Figure 1. — Fmn2 association with ER at spindle periphery and local actin polymerization. (A) Timeline of events in an unperturbed MI mouse oocyte. PBE, polar body extrusion. (B) Localization of Fmn2-AcGFP before and after GVBD (time stamps according to Video 1). Red, chromosomes. (C) Colocalization of Fmn2 with ER stained with blue-white DPX. Arrows point to some distinct structures with colocalization. (D) Overexpression of Sec61-β–mKate2 induces ER tubularization and accumulation of Fmn2 and F-actin along tubular ER. (E) Nocodazole treatment at GVBD + 0.5 h abolished normal Fmn2 distribution and actin enrichment around the chromosomes. (F) Plots of SEM and mean Pearson coefficient (n = 3) as a function of spatial shift in units of micrometers from spatial correlation analysis. (G) Percentage (mean and SEM from three experiments) of oocytes succeeded in chromosome migration when overexpressing Sec61-β–mKate2 (as in D; no Sec61-β overexpression [OE], n = 57; Sec61-β overexpression, n = 71) or treated with nocodazole at GVBD + 0.5 h to block ER congregation (as in E; no nocodazole [Noco], n = 59; nocodazole, n = 65). P < 0.005. (H) F-actin in the MI oocyte (GVBD + 6 h) visualized by strong expression of UtrCH-GFP (left) or microinjection of trace fluorescent phalloidin (right). A magnified view of the boxed region on the left is shown in the middle. (I) Staining of fixed oocytes (GVBD + 6 h) with Alexa Fluor 633–labeled phalloidin. (J) Right three images are time projections of a difference video with chromatin staining (blue) from the box of the UtrCH-GFP image on the left. (K) Montage of new F-actin growth near chromosomes after cytochalasin D washout. Full view is shown in Fig. S1 J. (L) Increase of intensity ratio of chromosome region (circled area; the data shown are from a representative video shown in Fig. S1 J, n = 5). Bars: (B–E, H [left and right], I, J [left], and L) 20 µm; (H [middle], J [right], and K) 10 µm.