SC Dynamics at Steady State Figure 5. A non-uniform spatial pattern of post-synapsis Zip1 deposition is observed independent of synapsis initiation components, Pch2, Pph3, and the presence of a homolog. (A) Meiotic surface-spread chromosomes labeled with DAPI to show DNA (white, left panels) anti-Zip1 (red), anti-GFP (labels Zip1-GFP, green) and Ctf19-MYC (labels centromeres, bluish purple) to show representative images of Zip1-GFP incorporation (after a 45 minute induction) into previously established SCs in mutants deficient in synapsis initiation components, and in haploid meiosis (K139). Scale, 1 mm. (B) Scatterplot displays the cumulative length of non-focal Zip1-GFP incorporation per mm of Zip1 in nuclei from various mutant strains, under induced (45 minutes) or uninduced conditions. Circles indicate values of Zip1-GFP length per total Zip1 length for individual nuclei; horizontal and error bars indicate mean and standard error of the mean. Complete genotypes of wt (K39), zip3 (K59), zip3 fpr3 (K83), zip4 zip3 fpr3 (K91), zip2 zip3 fpr3 (K92), fpr3 (K84), pph3 (AM2560) are listed in Table S1. An analogous graph is depicted below for wild type and pch2 (K117) mutant strains; these strains were analyzed independently of the experiments pictured above. All experiments were completed at least twice. (A scatterplot showing the extent of Zip1 assembled in the meiotic nuclei of these various strains at the time of ZIP1-GFP induction is shown in Figure S4.). doi:10.1371/journal.pgen.1002993.g005 long stretch of Zip1-GFP, while 30% of centromeres lacked Zip1- GFP staining (Figure 4C). These data suggest that, centromeres are not necessarily preferred over chromosomal arm sites for initial post-synapsis incorporation of Zip1. Sites of Initial Zip1 Incorporation into Previously Established SC Do Not Strictly Reflect Pattern of Zip1 ‘‘Peaks’’ in Full SC Previous work had suggested a non-uniformity in the structure of the budding yeast SC central region: discrete, local domains of SC central region stain more intensely with Zip1 antibody (Zip1 ‘‘peaks’’) as compared to other domains (‘‘valleys’’) [30]. Using a subset of immuno-stained meiotic nuclei that were maximally surface-spread such that Zip1 staining formed visually-apparent domains of thicker (peaks) and thinner (valleys) Zip1 staining regions along the length of the SC, we explored whether early sites of post-synapsis Zip1-GFP incorporation strictly associate with either ‘‘peaks’’ or ‘‘valleys’’ of a previously deposited SC. We observed a substantial fraction of post-synapsis Zip1-GFP incor- poration events (25% (n = 300)) that clearly colocalized with a ‘‘valley’’ of SC central region (Figure S3). The remaining sites either localized at a recognizable ‘‘peak’’ of SC central region (45%) or could not be unambiguously assigned to either an SC ‘‘peak’’ or ‘‘valley’’ (30%). Thus, the distribution of initial Zip1- GFP incorporation sites along the length of the SC does not precisely mimic the global domain structure of Zip1 within the SC itself. This conclusion is further supported by our observation that PLOS Genetics | www.plosgenetics.org 8 October 2012 | Volume 8 | Issue 10 | e1002993