DNA Topoisomerase II Is Not Detectable on Lampbrush Chromosomes but Enriched in the Amplified Nucleoli of Xenopus Oocytes

Abstract

In somatic cells DNA topoisomerase 11 (topo 11) is thought to be involved in the domain organization of the genome by anchoring the basis of chromatin loops to a chromosomal scaffold. Lampbrush chromosomes of amphibian oocytes directly display this radial loop organization in cytological preparations. In order to find out whether topo 11 may play a role in the organization of these meiotic chromosomes, we performed immunofluorescence studies using antibodies against Xenopus topo 11. Our results indicate that topo 11 is apparently absent from lampbrush chromosomes and is hence unlikely to act as a "fastener" of the numerous lateral chromosomalloops. Topo 11 was, however, enriched in the amplified nucleoli of Xenopus oocytes. © 1993 Academic Press, Inc. INTRODUCTIONThe chromatin fiber of eukaryotic cells is folded into a series of loops or domains which are stabilized by a structural framework called nuclear matrix or chromosomal scaffold. The loops are thought to be anchored at their base to the matrix or scaffold via specific attachment sites, the so-called matrix or scaffold attachment regions (MARs or SARs). Loop length is variable, ranging from 5 to 200 kbp of DNA [for reviews see 1, 2]. Unfortunately, the chromatin loop structures cannot be directly visualized in interphase nuclei with cytological methods because the chromatin strands are highly entangled.The situation is different for amphibian lampbrush chromosomes which can be readily examined under almost native conditions by light microscopy [3]. The structural organization of these meiotic chromosomes exhibits a striking similarity to the loop model described above. Each lampbrush chromosome consists of a chromosomal axis from which numerous loops radiate laterally. The axis represents a linear array of chromo-1 To whom correspondence and reprint requests should be addressed at Zoologie I, Biozentrum Am Hubland, D-97074 Wiirzburg, Germany. Fax: 0049 931 888 4252. 255 meres containing highly condensed chromatin, while the lateral loops consist of decondensed, transcriptionally active chromatin covered with numerous nascent transcripts. Because lampbrush chromosome architecture resembles so closely the proposed organization of somatic interphase chromatin and mitotic chromosomes, it is tempting to speculate that the same molecular components might be involved in the maintenance of chromatin loops.One of the best characterized components of the chromosomal scaffold is DNA topoisomerase 11 (topo 11) [4][5][6][7]. Besides its enzymatic activity that creates transient double-strand breaks in duplex DNA and passes DNA strands through one another, the protein has been shown to interact with the MAR and SAR elements [8]. Thus, topo 11 is thought to have catalytic as well as structural functions. Topo 11 is not only associated with the scaffold of mitotic chromosomes but also the cores and centromeric regions of spermatocyte-derived meiotic chromosomes [9]. Recently Luke and Bogenhagen identified by biochemical means topo 11 in the germinal vesicles of Xenopus o...

Centromeric protein bodies on avian lampbrush chromosomes contain a protein detectable with an antibody against DNA topoisomerase II

“…Thus, it is reasonable to expect topo II to be involved in the process of active pol II transcription on the lateral loops of the LBCs. However, immunofluorescence analysis did not reveal direct association of topo II with lateral loops in avian (our data) or in amphibian GVs (Fisher et al 1993;Hock et al 1996). Immunoelectron microscopy studies performed on thin sections of chaffinch GVs showed the presence of topo II in the nucleoplasm.…”

“…Topoisomerase II was not detectable in other parts of the LBCs (Fisher et al 1993). Because of the morphological resemblance between avian PBs and amphibian axial granules, we used mAb 4A6 to stain GV spreads from both chaffinch and pigeon previtellogenic oocytes.…”

Subnuclear localization of S/MAR-binding proteins is differently affected by in vitro stabilization with heat or Cu 2+

“…As for DNA topoisomerase II α, its role as a structural link anchoring the bases of chromatin loops to the nuclear matrix or chromosome scaffold in mammalian cells has been repeatedly proposed (see for example Cockerill and Garrard 1986;Gasser and Laemmli 1986;Adachi et al 1989), even though more recent evidence collected from Xenopus oocytes has challenged such a hypothesis (Fischer et al 1993;Hirano and Mitchison 1993).…”

Threonine 1342 in Human Topoisomerase IIα Is Phosphorylated Throughout the Cell Cycle

“…In addition to catalytic activity, topoisomerase II may function to anchor chromosomal DNA loops to the nuclear scaffold (8,9); topoisomerase II is a major non-histone protein present in nuclear scaffold fractions (10,11), and DNA sequences that bind preferentially to the nuclear matrix/chromosomal scaffold (MAR/SAR) contain topoisomerase II cleavage consensus (12). In contrast, the role of topoisomerase II in the maintenance of chromosomes was not supported in studies done using Xenopus egg extracts or Xenopus embryos (13,14). Topoisomerase II has been considered as one of the targets for anticancer drugs (15).…”