Sister chromatid cohesion is maintained from DNA replication to metaphase-to-anaphase transition by multisubunit protein complexes called cohesin, which include at least four proteins, SMC1alpha, SMC3, Rad21 and either SA1 or SA2, in mammalian somatic cells. We report here the first evidence of the involvement of Rec8 protein, a mammalian homolog of yeast Rec8p, in meiosis-specific chromosome behavior in mammals. In immunoblotting and immunohistochemical analysis using specific antibodies against mouse Rec8, we found that Rec8 was expressed in the testis but not in the kidney or liver; more precisely, it was expressed in spermatocytes and spermatids but not in spermatogonia or other somatic cells. We also found that Rec8 is present in both phosphorylated and dephosphorylated states in vivo. Immunoprecipitation analyses revealed that Rec8 associates with other cohesin proteins, SMC1beta (meiosis-specific protein) and SMC3 and with a component of synaptonemal complexes, SCP3, but not with SMC1alpha. In meiotic chromosome spreads, Rec8 was localized along the axial/lateral elements of the synaptonemal complexes in meiotic prophase from the leptotene to diplotene stages. At later stages, diakinesis and metaphase I, Rec8 was localized along the interstitial axes of chromosomes, including both centromere and arm regions of chromosomes. However, concomitantly with separation of homologous chromosomes in anaphase I, Rec8 was no longer detected along the arm regions, while it persisted on centromere regions up to metaphase II. In anaphase II, the centromeric signals were diminished. We propose from these results that mammalian Rec8 protein, in association with SMC3 and SMC1beta but not SMC1alpha, is involved in meiosis-specific chromosome behavior, and that homologous chromosome separation is triggered by selective loss of Rec8 from chromosome arms in meiosis I, while sister chromatid cohesion is maintained until metaphase II/anaphase II transition by centromeric Rec8 during mammalian meiosis.