Fig. 2: dHJ–ZMM protein interplay maintains chromosome synapsis and protects crossover precursors.
From: Holliday junction–ZMM protein feedback enables meiotic crossover assurance
a, Representative chromosome spreads at the indicated times in SPM, after Yen1ON induction by β-oestradiol addition (or methanol control) at 7 h, immunostained for Ecm11–Gmc2 (green) and Zip3 (magenta). The arrowheads mark Zip3 localizing to the Ecm11–Gmc2 polycomplex. b, Quantification of Zip3 focus number from a. Data are mean ± s.d. n = 30 nuclei per timepoint. Statistical analysis was performed using Kruskal–Wallis tests with Dunn’s multiple-comparison test (P ≤ 0.001). Representative of two biological replicates. c, The experimental set-up for conditional depletion of a ZMM protein in pachytene-arrested ndt80∆ cells. Os, Oryza sativa. d, Western blot analysis of Zip3AID levels in cells at the indicated times in SPM, treated as described in c. Crm1 was used as the protein loading control. The asterisks indicate putative SUMOylated Zip3. Representative of two biological replicates. e, Representative images of chromosome spreads from d, immunostained for Zip1 (green) and Zip3AID (magenta). ZIP3AID cells exhibit frequent polycomplex formation but show normal spore viability (Supplementary Table 2). f, Quantification of Zip1 synapsis and polycomplexes from e. n = 50 nuclei per timepoint, representative of two biological replicates. g,h, Southern blot (g) and quantification of joint molecules and non-crossover (NCO1) and crossover (CO2) products (h) at the HIS4::LEU2 recombination hotspot from d. mcJM, multichromatid DNA joint molecule; P1, parental 1; P2, parental 2. i, The experimental set-up for conditional depletion of a ZMM protein and Sgs1 in pachytene-arrested ndt80∆ cells. j, Western blot analysis of Zip3AID and Sgs1AID levels as in d for ZIP3AIDSGS1AID cells treated as described in i. Representative of two biological replicates. k,l, Southern blot (k) and quantification of joint molecules and non-crossover and crossover products (l) as in g and h, respectively, for the experiment in j. m, Representative images of chromosome spreads as in e, for the experiment in j. Both Zip3AID and Sgs1AID carry the Myc epitope tag. n, Quantification of Zip1 synapsis and polycomplexes as in f, for the experiment in m. n = 50 nuclei per timepoint, representative of two biological replicates. For a, e and m, scale bars, 2 μm.
