Chwen-Huey Wu

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The histone variant H2AZ is incorporated preferentially at specific locations in chromatin to modulate chromosome functions. In Saccharomyces cerevisiae, deposition of histone H2AZ is mediated by the multiprotein SWR1 complex, which catalyzes ATP-dependent exchange of nucleosomal histone H2A for H2AZ. Here, we define interactions between SWR1 components and(More)
NELF and DSIF collaborate to inhibit elongation by RNA polymerase IIa in extracts from human cells. A multifaceted approach was taken to investigate the potential role of these factors in promoter proximal pausing on the hsp70 gene in Drosophila. Immunodepletion of DSIF from a Drosophila nuclear extract reduced the level of polymerase that paused in the(More)
Variant histone H2AZ-containing nucleosomes are involved in the regulation of gene expression. In Saccharomyces cerevisiae, chromatin deposition of histone H2AZ is mediated by the fourteen-subunit SWR1 complex, which catalyzes ATP-dependent exchange of nucleosomal histone H2A for H2AZ. Previous work defined the role of seven SWR1 subunits (Swr1 ATPase,(More)
Replication forks formed during rolling-circle DNA synthesis supported by a tailed form II DNA substrate in the presence of the primosome, the single-stranded DNA binding protein, and the DNA polymerase III holoenzyme (Pol III HE) that had been reconstituted from the purified subunits, beta, tau, and the gamma.delta complex, at limiting (with respect to(More)
The coordinated action of many enzymatic activities is required at the DNA replication fork to ensure the error-free, efficient, and simultaneous synthesis of the leading and lagging strands of DNA. In order to define the essential protein-protein interactions and model the regulatory pathways that control Okazaki fragment synthesis, we have reconstituted(More)
Individually purified subunits have been used to reconstitute the action of the Escherichia coli DNA polymerase III holoenzyme (Pol III HE) at a replication fork formed in the presence of the primosome, the single-stranded DNA binding protein, and a tailed form II DNA template. Complete activity, indistinguishable from that of the intact DNA Pol III HE,(More)
Studies with a rolling-circle DNA replication system reconstituted in vitro with a tailed form II DNA template, the DNA polymerase III holoenzyme (Pol III HE), the Escherichia coli single-stranded DNA binding protein, and the primosome, showed that within the context of a replication fork, the oligoribonucleotide primers that were formed were limited to a(More)
To investigate the role of the priming apparatus at the replication fork in determining Okazaki fragment size, the products of primer synthesis generated in vitro during rolling-circle DNA replication catalyzed by the DNA polymerase III holoenzyme, the single-stranded DNA binding protein, and the primosome on a tailed form II DNA template were isolated and(More)
NELF and DSIF act together to inhibit transcription elongation in vitro, and are implicated in causing promoter proximal pausing on the hsp70 gene in Drosophila. Here, further characterization of Drosophila NELF is provided. Drosophila NELF has four subunits similar to subunits of human NELF. The amino acid sequences of NELF-B and NELF-D are highly(More)
Genetic and proteomic approaches have identified numerous proteins that are potentially involved in regulating transcriptional elongation, but the mechanisms of action of these proteins remain largely unknown. We describe an experimental approach using native gel electrophoresis for studying interactions of elongation factors with isolated Pol II elongation(More)
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