Figure 3.

Phosphorylation of Multiple LMN-1 Residues Is Required for Lamina Remodeling and Efficient Chromosome End Movement in Early Meiosis but Dispensable for Mitosis

(A) Representative still images of the first mitotic division in early embryos: histone:mCherry marks mitotic chromosomes; GFP::LMN-1 indicates LMN-1. Note the lack of complete lamina disassembly during late metaphase and anaphase, with LMN-1 localizing to the remnants of the nuclear envelope separating maternal and paternal chromosome masses (arrowheads). Nevertheless, embryos are fully viable (see Table S1). For quantifications, see Figure S3A. Scale bars, 10 μm.

(B) Localization of GFP::LMN-1 in the distal germline of untreated and detergent-treated [gfp::lmn-1^S8A] gonads. In this mutant, all phosphorylation sites flanking the coiled-coil domain have been changed to non-phosphorylatable residues. Arrows indicate the onset of meiosis. Scale bars, 10 μm. Gonads shown in (B) consist of multiple maximum projection images stitched together to show larger sections of the gonads.

(C) GFP::LMN-1 intensity in detergent-treated gonads of the wild-type, [gfp::lmn-1^S32,403A], and [gfp::lmn-1^S8A] mutants: the meiotic nuclear signal is presented as a percentage of the nuclear signal in the mitotic zone. [gfp::lmn-1], n = 5 gonads (at least 20 mitotic zone and 8 meiotic nuclei analyzed in each gonad); [gfp::lmn-1^S32,403A], n = 5 gonads (at least 22 mitotic zone and 9 meiotic nuclei analyzed in each gonad); [gfp::lmn-1^S8A], n = 5 gonads (at least 15 mitotic zone and 10 meiotic nuclei analyzed in each gonad). Non-significant differences (p > 0.05) are not indicated. Error bars represent SEM.

(D) GFP::LMN-1 intensity in mitotic zone (MZ) and transition zone (TZ) nuclei of the wild-type and the two non-phosphorylatable lmn-1 mutants: ratio of signals at the nuclear periphery/nuclear interior. [gfp::lmn-1], n = 16 nuclei in each zone; [gfp::lmn-1^S32,403A], n = 16 nuclei in each zone; [gfp::lmn-1^S8A], n = 16 nuclei in each zone. For all genotypes analyzed nuclei were taken from four different gonads. p values were calculated using the two-tailed Student's t test. Error bars represent SEM.

(E) Tracking of individual SUN-1::mRuby aggregates in transition zone nuclei of worms expressing wild-type or non-phosphorylatable LMN-1. Upper panel: the speed of individual SUN-1 aggregates from 14 nuclei/genotype is shown (wild-type, n = 207 aggregates; [gfp::lmn-1^S8A], n = 178 aggregates). Lower panel: projected displacement tracks for SUN-1 aggregates in representative nuclei of both genotypes. p value was calculated using the Mann-Whitney U test. Scatter plots indicate the mean and SD. For additional examples, see Figure S3B. Scale bars, 5 μm.

(F) Tracking of individual HIM-8::mCherry aggregates in early transition zone nuclei of worms expressing wild-type (n = 15 nuclei) and non-phosphorylatable LMN-1 in the sun-1(jf18) mutant ([gfp::lmn-1^S32,403A], n = 7 nuclei; [gfp::lmn-1^S8A], n = 10 nuclei). In this mutant, the cytoskeletal forces responsible for directed chromosome movement are eliminated. Background Brownian motion was assessed in worms mounted in 10% Na azide (n = 8 nuclei). p values were calculated using the two-tailed Student's t test. Scatter plots indicate the mean and SD.

^∗p < 0.05, ^∗∗p < 0.01, ^∗∗∗p < 0.001.