DNA topoisomerase II (topo II) catalyzes a strand passage reaction in that one duplex is passed through a transient brake or gate in another. Completion of late stages of neuronal development depends on the presence of active β isoform (topo IIβ). The enzyme appears to aid the transcriptional induction of a limited number of genes essential for neuronal maturation. However, this selectivity and underlying molecular mechanism remains unknown. Here we show a strong correlation between the genomic location of topo IIβ action sites and the genes it regulates. These genes, termed group A1, are functionally biased towards membrane proteins with ion channel, transporter, or receptor activities. Significant proportions of them encode long transcripts and are juxtaposed to a long AT-rich intergenic region (termed LAIR). We mapped genomic sites directly targeted by topo IIβ using a functional immunoprecipitation strategy. These sites can be classified into two distinct classes with discrete local GC contents. One of the classes, termed c2, appears to involve a strand passage event between distant segments of genomic DNA. The c2 sites are concentrated both in A1 gene boundaries and the adjacent LAIR, suggesting a direct link between the action sites and the transcriptional activation. A higher-order chromatin structure associated with AT richness and gene poorness is likely to serve as a silencer of gene expression, which is abrogated by topo IIβ releasing nearby genes from repression. Positioning of these genes and their control machinery may have developed recently in vertebrate evolution to support higher functions of central nervous system.

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Nucleosome assembly protein 1 (Nap1) is widely conserved from yeasts to humans and facilitates nucleosome formation in vitro as a histone chaperone. Nap1 is generally localized in the cytoplasm, except that subcellular localization of Drosophila melanogaster Nap1 is dynamically regulated between the cytoplasm and nucleus during early development. The cytoplasmic localization of Nap1 is seemingly incompatible with the proposed role of Nap1 in nucleosome formation, which should occur in the nucleus. Here, we have examined the roles of a putative nuclear export signal (NES) sequence in yeast Nap1 (yNap1). yNap1 mutants lacking the NES-like sequence were localized predominantly in the nucleus. Deletion of NAP1 in cells harboring a single mitotic cyclin gene is known to cause mitotic delay and temperature-sensitive growth. A wild-type NAP1 complemented these phenotypes while nap1 mutant genes lacking the NES-like sequence or carboxy-terminal region did not. These and other results suggest that yNap1 is a nucleocytoplasmic shuttling protein and that its shuttling is important for yNap1 function during mitotic progression. This study also provides a possible explanation for Nap1's involvement in nucleosome assembly and/or remodeling in the nucleus.Chromatin is one of the hallmarks of eukaryotes. The eukaryotic genome DNA is complexed with chromosomal proteins to form the chromatin structure in the nucleus. Nuclear reactions such as DNA replication, transcription, DNA repair, and recombination take place on the chromatin and are regulated by the disruption and assembly of chromatin, namely chromatin remodeling. Recently, a variety of chromatin remodeling factors has been identified, including histone modification enzymes and ATP-dependent factors (reviewed in references 1, 2, 22, and 42). The histone chaperone family is one of the chromatin remodeling factors that binds to core histones and facilitates assembly and remodeling of the chromatin in an ATP-independent manner. This family includes various kinds of proteins, such as nucleoplasmin (31) and N1/N2 (28) in Xenopus, nucleosome assembly protein 1 (designated Nap1 in this paper) and Nap2 (13, 48), template-activating factor I␣

Template activating factor I (TAF-I) was originally identified as a host factor required for DNA replication and transcription of adenovirus genome complexed with viral basic proteins. Purified TAF-I was shown to bind to core histones and stimulate transcription from nucleosomal templates. Human TAF-I consists of two acidic proteins, TAF-I␣ and TAF-I␤, which differ from each other only in their amino-terminal regions. Here, we report that TAF-I decondenses demembraned Xenopus sperm chromatin. Human TAF-I␤ has a chromatin decondensation activity comparable to that of NAP-I, another histone binding protein, whereas TAF-I␣ has only a weak activity. Analysis of molecular mechanisms underlying the chromatin decondensation by TAF-I revealed that TAF-I interacts directly with sperm basic proteins. Deletion of the TAF-I carboxyl-terminal acidic region abolishes the decondensation activity. Interestingly, the acidic region itself is not sufficient for decondensation, since an amino acid substitution mutant in the dimerization domain of TAF-I which has the intact acidic region does not support chromatin decondensation. We detected the ␤ form of TAF-I in Xenopus oocytes and eggs by immunoblotting, and the cloning of its cDNA led us to conclude that Xenopus TAF-I␤ also decondenses sperm chromatin. These results suggest that TAF-I plays a role in remodeling higher-order chromatin structure as well as nucleosomal structure through direct interaction with chromatin basic proteins.