During meiosis, chromosomes assemble specialized structures that promote: 1) establishment and maintenance of homolog pairing, 2) crossing over between homologs and 3) segregation of homologs at the meiosis I division. We are taking several approaches to investigate the dynamic nature of these structures and how their assembly is regulated and coordinated with homolog pairing. During meiotic prophase, axial elements are assembled along the lengths of chromosomes and axes of aligned homologs are connected by the synaptonemal complex (SC). Axial elements are built on a foundation of the cohesin complex, made up of SMC-1, SMC-3, REC-8 and SCC-3. Whereas previous analysis (Chan et al. 2003) had suggested that SMC subunits can load independently of non-SMC subunits, we find that SMC-1 does not localize to chromosomes in scc-3
) mutants, implying that SMC loading does require SCC-3. Instead SMC-1 is present in aggregates that also contain SC components SYP-1 and HIM-3. These aggregates are lost in syp-1 scc-3
double mutants but chromosomal localization of SMC-1 is not restored, implying that SCC-3 plays a positive role in chromosomal loading of cohesin and does not function solely to inhibit premature associations of SC components and cohesin subunits. Our results indicate that SCC-3 is involved in the proper chromosomal assembly of cohesin and SC. Prior to SC assembly, homolog pairing is accompanied by nuclear reorganization of chromosomes into a clustered configuration. Such nuclear reorganization is absent in pairing-defective hal-2
mutants. In addition to defective pairing, hal-2
mutants load SYP-1 incorrectly on unpaired homologs. Pairing at the pairing centers is substantially restored in hal-2
double mutants, implying that incorrect loading of SYP proteins is partially responsible for inhibiting homolog pairing. hal-2
mutants lack the extended region of clustered chromosome configuration seen in syp mutants, however, implying that HAL-2 has additional roles in promoting normal chromosome organization beyond inhibiting association of SYP proteins with unpaired homologs. Thus, we have identified HAL-2 as a novel component of the meiotic machinery involved in coordination of early meiotic events. While EM images give an impression of the SC as a static scaffold-like structure, recent findings suggest that the SC is more dynamic than previously thought. We will conduct FRAP analysis to investigate SC protein dynamics, using strains expressing GFP- and mCherry-tagged SC components that correctly localize to chromosomes.