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The fly Drosophila melanogaster is one of the most intensively studied organisms in biology and serves as a model system for the investigation of many developmental and cellular processes common to higher eukaryotes, including humans. We have determined the nucleotide sequence of nearly all of the approximately 120-megabase euchromatic portion of the(More)
The protein Cdc45 plays a critical but poorly understood role in the initiation and elongation stages of eukaryotic DNA replication. To study Cdc45's function in DNA replication, we purified Cdc45 protein from Drosophila embryo extracts by a combination of traditional and immunoaffinity chromatography steps and found that the protein exists in a stable,(More)
The Drosophila Myb complex has roles in both activating and repressing developmentally regulated DNA replication. To further understand biochemically the functions of the Myb complex, we fractionated Drosophila embryo extracts relying upon affinity chromatography. We found that E2F2, DP, RBF1, RBF2, and the Drosophila homolog of LIN-52, a class B synthetic(More)
The origin recognition complex (ORC) is required to initiate eukaryotic DNA replication and also engages in transcriptional silencing in S. cerevisiae. We observed a striking preferential but not exclusive association of Drosophila ORC2 with heterochromatin on interphase and mitotic chromosomes. HP1, a heterochromatin-localized protein required for position(More)
MCM2-7 proteins provide essential helicase functions in eukaryotes at chromosomal DNA replication forks. During the G1 phase of the cell cycle, they remain loaded on DNA but are inactive. We have used recombinant methods to show that the Drosophila MCM2-7 helicase is activated in complex with Cdc45 and the four GINS proteins (CMG complex). Biochemical(More)
Myb-MuvB (MMB)/dREAM is a nine-subunit complex first described in Drosophila as a repressor of transcription, dependent on E2F2 and the RBFs. Myb, an integral member of MMB, curiously plays no role in the silencing of the test genes previously analyzed. Moreover, Myb plays an activating role in DNA replication in Drosophila egg chamber follicle cells. The(More)
Initiation of cellular DNA replication is tightly controlled to sustain genomic integrity. In eukaryotes, the heterohexameric origin recognition complex (ORC) is essential for coordinating replication onset. Here we describe the crystal structure of Drosophila ORC at 3.5 Å resolution, showing that the 270 kilodalton initiator core complex comprises a(More)
There is considerable interest in the developmental, temporal and tissue-specific patterns of DNA replication in metazoans. Site-specific DNA replication at the chorion loci in Drosophila follicle cells leads to extensive gene amplification, and the organization of the cis-acting DNA elements that regulate this process may provide a model for how such(More)
Drosophila origin recognition complex (ORC) localizes to defined positions on chromosomes, and in follicle cells the chorion gene amplification loci are well-studied examples. However, the mechanism of specific localization is not known. We have studied the DNA binding of DmORC to investigate the cis-requirements for DmORC:DNA interaction. DmORC displays at(More)
Gene amplification at the chorion loci in Drosophila ovarian follicle cells is a model for the developmental regulation of DNA replication. Previously, we showed that the Drosophila homolog of the Myb oncoprotein family (DmMyb) is tightly associated with four additional proteins and that DmMyb is required for this replication-mediated amplification. Here we(More)