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Bloom's syndrome (BS) is an autosomal disorder characterized by predisposition to a wide variety of cancers. The gene product whose mutation leads to BS is the RecQ family helicase BLM, which forms a complex with DNA topoisomerase III␣ (Top3␣). However, the physiological relevance of the interaction between BLM and Top3␣ within the cell remains unclear. We show here that Top3␣ depletion causes accumulation of cells in G 2 phase, enlargement of nuclei, and chromosome gaps and breaks that occur at the same position in sister chromatids. The transition from metaphase to anaphase is also inhibited. All of these phenomena except cell lethality are suppressed by BLM gene disruption. Taken together with the biochemical properties of BLM and Top3␣, these data indicate that BLM and Top3␣ execute the dissolution of sister chromatids.Eukaryotic TOP3 was first identified in Saccharomyces cerevisiae as a gene that is required to suppress recombination between repeated sequences (28). Deletion of TOP3 results in a slow-growth phenotype that is suppressed by the disruption of SGS1, the gene encoding the sole RecQ helicase in S. cerevisiae (5). Further analyses revealed that the function of Sgs1 is closely associated with that of DNA topoisomerase III (Top3) (2,13,19,27). The close relationship between RecQ helicases and Top3 seems to be maintained in higher eukaryotes. Higher eukaryotic cells have two Top3s, Top3␣ and Top3␤ (8,22,23). Knocking out the Top3␣ gene in mice results in embryonic lethality (17), while knocking out Top3␤ does not affect development but reduces the life span (15). Various Top3 and RecQ helicase molecules have been reported to interact physically, including Top3␣ and BLM (35), one of the RecQ family helicases in higher eukaryotic cells (3). BLM is a causative gene for Bloom's syndrome (3), which is an autosomal disorder characterized by predisposition to a wide variety of cancers (6). Biochemical analyses have suggested that BLM and Top3␣ together affect the in vitro resolution of a recombination intermediate containing a double Holliday junction (HJ) via a double-junction dissolution mechanism (34). However, the phenotypes of cells that lack Top3␣ have not been characterized precisely, since TOP3␣ knockout is lethal. Furthermore, the phenotypes of Top3␣-depleted cells before they die have not been examined. Moreover, the physiological relevance of the interaction between BLM and Top3␣ within the cell remains unclear. Therefore, elucidating higher eukaryotic Top3␣ function may enhance our understanding of the physiological roles of BLM.In this study, to assess the function of Top3␣ and its interactions with BLM, we constructed cells whose expression of Top3␣ can be switched off by doxycycline hydrochloride (Dox) treatment. To our knowledge, we present the first evidence to support the hypothesis that vertebrate Top3␣ together with the BLM helicase executes the dissolution of sister chromatids during DNA replication. MATERIALS AND METHODSPlasmid construction. Fragments of chicken TOP3␣ and TOP3␤ cDNAs ...

Rat heme-binding protein 23 (HBP23)/peroxiredoxin (Prx I) belongs to the 2-Cys peroxiredoxin type I family and exhibits peroxidase activity coupled with reduced thioredoxin (Trx) as an electron donor. We analyzed the dimer-oligomer interconversion of wild-type and mutant HBP23/Prx I by gel filtration and found that the C52S and C173S mutants existed mostly as decamers, whereas the wild type was a mixture of various forms, favoring the decamer at higher protein concentration and lower ionic salt concentration and in the presence of dithiothreitol. The C83S mutant was predominantly dimeric, in agreement with a previous crystallographic analysis (Hirotsu, S., Abe, Y., Okada, K., Nagahara, N., Hori, H., Nishino, T., and Hakoshima, T.

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Molecular cloning experiments indicated that BW10KD as a BW allergen was a member of the 2S-albumin multigene family.