Author Summary Sexually reproducing parents use meiosis to generate specialized cells in which chromosome sets are reduced from two to one. Accurate chromosome reduction relies on the prior establishment of pair-wise associations between homologous chromosomes (homologs); mainte- nance of paired associations typically occurs via inter- homolog crossover recombination events. The proteina- ceous, structurally conserved synaptonemal complex (SC) assembles along the full length of aligned homolog axes. The SC is the context in which crossover recombination events mature, and it influences both the level and distribution of crossover events between homologs. However it is not clear whether the SC maintains the capacity to remodel, for example to structurally accom- modate recombination events. We explore the dynamics of budding yeast SC and find that full-length SCs exhibit ongoing subunit incorporation but little subunit turnover during a meiotic cell cycle arrest, thus SC grows over time. Interestingly, initial subunit incorporation into full-length SCs occurs predominantly at or adjacent to Zip3 foci, a presumed marker of crossover sites. Our observations suggest that budding yeast SC continues to assemble during a steady state and that, while it may have little capacity for global turnover after installation, the SC maintains differential assembly dynamics at recombina- tion-associated perturbations in the meiotic axis. SC may be required for a normal number and distribution of crossover recombination events (which will in turn ensure the persistence of homologous associations after SC disassembly and until chromosome segregation on the meiosis I spindle) [2–8]. Zip1 is a primary structural component of the SC central region in budding yeast [9–11]. Like most SC central region proteins identified to date, Zip1 contains an extensive coiled-coil domain flanked by globular ends, and is predicted to form dimers. The structure of SC central region proteins resembles that of intermediate filament subunits and suggests a capacity to self- assemble [3,12]. Elegant immuno-electron microscopy experi- ments in budding yeast demonstrated that Zip1 subunits interact with one another near their amino termini, and interact with chromosome cores at their carboxyl terminal ends [9]. Additional proteins that do not share structural similarity to typical transverse filament proteins may also contribute to establishing SC central region. For example, the Small Ubiquitin-like MOdifier protein, SUMO, has been implicated in SC central region assembly on the basis of a dependence on central region proteins for its localization to SC [12–15]. The Synapsis Initiation Complex (SIC) proteins Zip2, Zip3 and Zip4 promote SC central region assembly from distinct synapsis initiation sites along the length of the chromosome [16–18]. A set of synapsis initiation sites corresponds to centromeres, while the remainder are presumed, based on the number and distribution of SIC foci along chromosome arms, to correspond to crossover recombination sites [19]. Interestingly, SIC proteins remain localized as discrete foci along chromosomes even after full-length SC has been deposited [16–18], raising the question of whether SICs have a later role in crossover recombination or SC maintenance at discrete sites, after their initial role in SC assembly. The existence of multiple layers of regulation that prevent inappropriate synapsis initiation suggests that SC may act as an irreversible tether between chromosomes [5]. Initial SC assembly interfaces closely with the homolog pairing process: Observations SC Dynamics at Steady State in multiple organisms of SC protein aggregation (polycomplex formation) in pairing-defective meiotic mutants indicate that cells normally regulate SC subunits such that their assembly on chromatin is contingent on homology verification [2,6,20], and this notion has been borne out by the identification of checkpoint- like pathways that prevent SC assembly when homologous pairing fails [5,21,22]. However, the possibility of inappropriate initiations or interlocks between partially synapsed chromosome pairs raises the question of whether the SC central region maintains a capacity to remodel. Furthermore, assembled SC is the context within which meiotic recombination events mature, and analyses of SC- deficient meiotic mutants suggest that the SC functionally interfaces with at least a subset of recombination events [17,18,23,24]. Whether the dynamics or composition of SC central region is altered at Spo11-mediated recombination sites is not known. Here we investigate the dynamics of the fully assembled SC in budding yeast meiotic cells that are arrested at the pachytene stage of meiotic prophase (when chromosomes are fully synapsed). Multimeric protein structures that exhibit both ongoing incorpo- ration and ongoing exit (‘‘treadmilling’’) might be rapidly disassembled by lowering the ‘‘on-rate’’ in a local region; perhaps the budding yeast SC structure exhibits such dynamics that would enable it to disassemble quickly. On the contrary, we demonstrate that Zip1 continually incorporates into full-length synaptonemal complex and that, globally, Zip1 does not exit to the same extent as it enters the SC structure in pachytene-arrested cells. The rate of SC assembly and maximum size of SC central region increases in direct proportion to Zip1 copy number. Our observations suggest that the budding yeast SC structure behaves more like a ‘‘tether’’ than a ‘‘treadmill’’ during pachytene arrest. Interestingly, fully assembled SCs exhibit a non-uniformity in SC assembly dynamics such that initial post-synapsis Zip1 incorporation is favored in the vicinity of recombination events. Results Full-Length Synaptonemal Complex Exhibits Ongoing Incorporation of Zip1 To explore the dynamics of the fully formed SC, we created a strain with inducible expression of the SC central region subunit, Zip1 (or a tagged version, Zip1-GFP, a kind gift of D. Kaback [25]), using the estrogen-regulated Gal4-ER transcription factor in trans [26,27]. In the absence of b-estradiol, chromosome spreads from meiotic cells in which ZIP1 or ZIP1-GFP is solely under GAL1 promoter control exhibited a zip1 phenotype: immunostaining of meiotic prophase chromosomes demonstrated that little to no Zip1 localizes to chromosomes in uninduced nuclei, and chromosomal axes often exhibited axial associations (intermittent points of contact presumed to be crossover recombination events, flanked by regions of abnormally loose axial alignment between homologs) (Figure S1). Moreover, in the absence of b-estradiol, the sporulation efficiency and spore viability of our ‘‘inducible-SC’’ strain phenocopied the zip1 (Figure S1). On the other hand, when sporulated in the presence of b- estradiol, cells carrying an inducible ZIP1 allele exhibited Zip1 or Zip1-GFP at the interface of aligned, homologous chromosome axes, and the sporulation efficiency as well as the viability of the spore products of these meiotic cells was rescued to wild-type levels (Figure S1). These data indicate that the estrogen-inducible Zip1 can functionally substitute for endogenous Zip1. Interestingly, when this strain was made homozygous for an ndt80 mutation, which arrests otherwise wild type cells at late prophase with fully synapsed chromosomes [28], we observed that Zip1 can assemble PLOS Genetics | www.plosgenetics.org 2 October 2012 | Volume 8 | Issue 10 | e1002993