Cloning and sequencing of cDNA encoding human DNA topoisomerase II and localization of the gene to chromosome region 17q21-22.

Abstract

Two overlapping cDNA clones encoding human DNA topoisomerase II were identified by two independent methods. In one, a human cDNA library in phage A was screened by hybridization with a mixed oligonucleotide probe encoding a stretch of seven amino acids found in yeast and Drosophila DNA topoisomerase II; in the other, a different human cDNA library in a Agtll expression vector was screened for the expression of antigenic determinants that are recognized by rabbit antibodies specific to human DNA topoisomerase II. The entire coding sequences of the human DNA topoisomerase II gene were determined from these and several additional clones, identified through the use of the cloned human TOP2 gene sequences as probes. Hybridization between the cloned sequences and mRNA and genomic DNA indicates that the human enzyme is encoded by a single-copy gene. The location of the gene was mapped to chromosome 17q21-22 by in situ hybridization of a cloned fragment to metaphase chromosomes and by hybridization analysis with a panel of mouse-human hybrid cell lines, each retaining a subset of human chromosomes.Eukaryotic DNA topoisomerase II is a ubiquitous ATPdependent type II topoisomerase (reviewed in refs. [1][2][3]

Bioflavonoids as Poisons of Human Topoisomerase IIα and IIβ

“…Many eukaryotic type II topoisomerases contain two Walker consensus ATP bindng motifs (105), Walker A and B (106)(107)(108). The Walker A site spans residues 161-166 (numbering is from human topoisomerase IIR) and is comprised of the G-X-X-G-X-G sequence (106)(107)(108)(109). The Walker B site, at residues 472-477, contains the G-X-G-X-X-G consensus (Table 1) (106)(107)(108)(109).…”

“…The Walker A site spans residues 161-166 (numbering is from human topoisomerase IIR) and is comprised of the G-X-X-G-X-G sequence (106)(107)(108)(109). The Walker B site, at residues 472-477, contains the G-X-G-X-X-G consensus (Table 1) (106)(107)(108)(109). The sequence at residues 161-166 is the site that actually is utilized for ATP binding in the eukaryotic type II enzyme (110).…”

Enhanced etoposide sensitivity following adenovirus-mediated human topoisomerase II α gene transfer is independent of topoisomerase II β

“…Then 20 µg of RNA was electrophoresed on a 1% formaldehyde/agarose gel and transferred to a Hybond-N + membrane (Amersham Corp., Arlington Heights, IL, USA). Human topo IIα gene probe ZII69 (Tsai-Pflugfeder et al, 1988), topo IIβ gene probe F12 (Austin et al, 1993;Herzog et al, 1998), and a GAPDH probe were used for hybridization. Probes were labelled using the Rediprime labelling system (Amersham).…”

Human Topoisomerase IIα is Phosphorylated in a Cell‐Cycle Phase‐Dependent Manner by a Proline‐Directed Kinase

“…In addition to this essential function during cell division, topoisomerase I1 enzymes play numerous other roles in DNA metabolism in eukaryotic cells, including mitotic chromosome condensation, relief of torsional stress and suppression of genomic instability (reviewed by Osheroff et al, 1991;Wang and Lynch, 1993;Watt and Hickson, 1994). Evidence has been presented that topoisomerase I1 performs a structural role in eukaryotic cell nuclei, anchoring topologically constrained domains of chromatin to the nuclear matrix or scaffold (reviewed by Roberge and Gasser, 1992).In contrast to lower eukaryotes, mammalian cells express two closely related, but genetically distinct, isozymes of topoisomerase I1 (Tsai-Pflugfelder et al, 1988;Drake et al, 1989;Chung et al, 1989;Tan et al, 1992;Jenkins et al, 1992;Austin et al, 1993), designated a (170-kDa form) and p (180-kDa form differential expression during the cell cycle and following oncogenic transformation (Woessner et al, 1990(Woessner et al, , 1991 that at least some functional specialization of these isozymes has evolved in mammalian cells. The extent to which the various functions of topoisomerase I1 are regulated by post-translational modification in human cells is unknown.…”

“…In contrast to lower eukaryotes, mammalian cells express two closely related, but genetically distinct, isozymes of topoisomerase I1 (Tsai-Pflugfelder et al, 1988;Drake et al, 1989;Chung et al, 1989;Tan et al, 1992;Jenkins et al, 1992;Austin et al, 1993), designated a (170-kDa form) and p . The roles of the topoisomerase IIa and topoisomerase IIp isozymes remain obscure, although it seems likely from their differential expression during the cell cycle and following oncogenic transformation (Woessner et al, 1990(Woessner et al, , 1991 that at least some functional specialization of these isozymes has evolved in mammalian cells.…”