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Almost a decade ago, Breuer and Pavan (1955) reported for the first time that during larval growth DNA synthesis proceeds disproportionately along polytene chromosomes of Rhynochosciara (R. angelae). In such chromosomes, treated by Feulgen reaction, the intensity of stain appeared heavier at the points where the bulbs were formed, thus indicating localized accumulation of DNA molecules. [...]

Disk performance for random access fares significantly worse compared to sequential access. Time required to transfer random blocks to or from disk is dominated by seeking and rotational delay. To improve the throughput and reduce the latency, one can apply techniques to increase the sequentiality of disk accesses, such as block rearrangement and replication. We introduce an approach to improve read performance by replicating blocks into file system free space at the block level. This makes the replication module independent of the file system and therefore easier to implement and verify. A solution that requires no changes to the file system is also easier to adopt. Supporting a new file system is a matter of writing a user-space component that understands its free block data structures. We implemented a prototype as a stacked device driver for Linux and evaluated its performance on a number of workloads.

Complete and accurate DNA replication is essential to maintain the genetic integrity in all organisms. In eukaryotes, the minichromosome maintenance (MCM) complex forms the catalytic core of the CMG helicase that unwinds DNA at the replication fork. We have previously identified a conserved MCM complex binding protein (MCM-BP) through a proteomic screen in human cells. In chapter two of this thesis, I show that MCM-BP makes an important contribution to nuclear morphology in human cells by affecting centrosome duplication. I also show that MCM-BP depletion results in G2 checkpoint signaling and the induction of replication stress. A recent study in Xenopus egg extracts has suggested that MCM-BP functions to unload the MCM complex from chromatin during S-phase. However, the mechanism of this process remains enigmatic. In chapter three of this thesis, I show that MCM-BP directly binds the de-ubiquitylating enzyme, USP7 and that this interaction is mediated by S158 on MCM-BP and the USP7 TRAF domain. Furthermore, I indicate a novel role for USP7 in DNA replication that involves unloading of the MCM complex during S-phase. Finally, my data suggest that MCM-BP tethers an interaction between the USP7 and the MCM complex to facilitate MCM complex unloading at the end of S-phase.

The Dbf4/Cdc7 kinase complex is required for the initiation of DNA replication and promotes this by acting upon members of the Mcm2-7 helicase. In addition to its role in replication, Dbf4/Cdc7 is a target of the S-phase checkpoint response through the Rad53 checkpoint kinase. In the budding yeast Saccharomyces cerevisiae, the regulatory subunit of this complex, Dbf4, is essential for kinase activity. Dbf4 is conserved throughout eukaryotes and contains three regions of discrete homology, termed the N, M, and C motifs, based on their location in the polypeptide chain. Motif C shows the highest conservation of all the motifs of Dbf4 and contains a CCHH type zinc finger. Mutation of the conserved cysteine and histidine residues of this zinc finger impair interactions with origin DNA and the Mcm2-7 helicase subunit Mcm2, but do not disrupt associations with Cdc7, Orc2, or Rad53. Cells where the endogenous Dbf4 CCHH zinc finger has been mutated exhibit slowed growth, and are delayed in their entry to, and progression through S-phase. These cells also display sensitivity upon long-term exposure to the ribonucleotide reductase inhibitor hydroxyurea (HU) and the DNA alkylating agent methyl methanesulfonate (MMS). The crystal structure of an amino-terminal region of Dbf4 containing motif N folds as a BRCA1-carboxy-terminal (BRCT) domain. This domain is required for the interaction with Rad53, but is not sufficient. A fragment of Dbf4 containing the BRCT domain and its fifteen preceding amino acids is sufficient to interact with Rad53 and folds as a modified BRCT domain containing an integral amino-terminal helical projection. Denoted the Helix-BRCT (HBRCT) domain, mutations that destabilize it abrogate the interaction with Rad53, and result in sensitivity to genotoxic agents. Dbf4 is recognized by the forkhead-associated FHA1 domain of Rad53, and the HBRCT domain of Dbf4 interacts directly with FHA1 in vitro. This interaction is phosphorylation independent and relies on a conserved lateral surface of FHA1, distinct from the phosphoepitope binding surface, which when mutated abrogates the interaction between Dbf4 and Rad53 and results in sensitivity to HU and MMS. The in vitro interaction between FHA1 and HBRCT does not require the ability of FHA1 to bind a phosphoepitope, while the in vivo interaction between full-length Rad53 and Dbf4 does. The FHA1 domain of Rad53 can simultaneously bind to a phosphopeptide and HBRCT, indicating that Rad53 recognition of Dbf4 may occur through a bipartite interaction using two surfaces of FHA1.

No description available.

Disk performance for random access fares significantly worse compared to sequential access. Time required to transfer random blocks to or from disk is dominated by seeking and rotational delay. To improve the throughput and reduce the latency, one can apply techniques to increase the sequentiality of disk accesses, such as block rearrangement and replication. We introduce an approach to improve read performance by replicating blocks into file system free space at the block level. This makes the replication module independent of the file system and therefore easier to implement and verify. A solution that requires no changes to the file system is also easier to adopt. Supporting a new file system is a matter of writing a user-space component that understands its free block data structures. We implemented a prototype as a stacked device driver for Linux and evaluated its performance on a number of workloads.

Thesis (Ph. D.)--University of Texas at Austin, 1998. / Vita. Includes bibliographical references (leaves 113-135). Available also in a digital version from Dissertation Abstracts.

Thesis (Ph.D.)--Aberdeen University, 2008. / Title from web page (viewed on Aug. 26, 2009). Includes bibliographical references.