Cell, Vol. 64, 137-146,


11, 1991, Copyright

0 1991 by Cell Press

Chromosome Assembly In Vitro: Topoisomerase II Is Required for Condensation Yasuhisa Adachi: May Luke,+ and Ulrich K. Laemmli’ * University of Geneva Departments of Biochemistry and Molecular Biology 1211 Geneva 4 Switzerland t Department of Pharmacological Sciences State University of New York at Stony Brook Stony Brook, New York 11794-8651

Summary The role of topoisomerase II (top0 II) in chromosome condensation was studied in a mitotic extract derived from Xenopus eggs by specific immunodepletion. HeLa nuclei, which have a high complement of endogenous topo II, are converted to mitotic chromosomes in the topo II-depleted extract equally well as in the control. Chicken erythrocyte nuclei, however, which have a very low content of topo II, do not convert to condensed chromosomes in the depleted extract, although their condensation is normal upon addition of purified topo II. Dosage experiments support the possible notion of a structural involvement of topo II in chromosome condensation. In the topo II-depleted extract the erythrocyte nuclei progress to precondensation chromosomes, which lack the nuclear membrane-lamina complex and consist of a cluster of swollen chromatids. Introduction The chromatin fiber of the eukaryotic chromosome is thought to be organized into topological loops, which are held together by a network of protein cross-ties that form the chromosomal scaffolding (e.g., Boy de la Tour and Laemmli, 1988). The major proteins of this scaffolding, postulated to be involved in the structural maintenance of metaphase chromosomes, are two high molecular weight proteins called SC1 and SC2 (Lewis and Laemmli, 1982). The protein SC1 was identified as topoisomerase II (top0 II). This enzyme partitions efficiently into the chromosomal scaffold (Earnshaw et al., 1985; Gasser el al., 1986) and also the nuclear matrix (Berrios et al., 1985). In line with the prediction of the chromosome loop model (Marsden and Laemmli, 1979), topo II immunolocalizes to a longitudinal, central region in expanded metaphase chromosomes (Earnshaw and Heck, 1985; Gasser et al., 1986). In more compact chromosomes, the scaffolding is helically folded with sister chromatids having predominantly opposite helical handedness (Boy de la Tour and LaemmIi, 1988). Topoisomerases have been implicated in several aspects of DNA metabolism (for recent reviews, see Wang, 1987; Sternglanz, 1989). Genetic analysis in yeast established that topo I or II can mutually substitute several of

their topological roles, but topo II has an essential function in the segregation of the intertwined daughter chromosomes at the end of DNA replication (Uemura and Yanagida, 1984; DiNardo et al., 1984; Holm el al., 1985). In yeast, genetic evidence also implicates topo II in chromosome condensation (Uemura et al., 1987), and drug inhibition studies in ceil-free nuclear/chromosome assembly systems suggest a role for topo II in chromatin condensation as well as decondensation (Newport, 1987; Newport and Spann, 1987). Topo II appears to be a marker for cell proliferation. During erythropoiesis in chicken, the topo II concentration drops from 7 x 104 copies per erythroblast to

Chromosome assembly in vitro: topoisomerase II is required for condensation.

The role of topoisomerase II (topo II) in chromosome condensation was studied in a mitotic extract derived from Xenopus eggs by specific immunodepleti...
5MB Sizes 0 Downloads 0 Views