Thesis (doctoral)--Lund University, 1997. / Added t.p. with thesis statement inserted.

Thesis (doctoral)--Lund University, 1997. / Added t.p. with thesis statement inserted.

The centromere is the site of kinetochore assembly that ensures proper chromosome segregation. Active centromeres are formed at chromosomal locations that do not appear to share homology between different species; this and other analyses has lead to the conclusion that centromeres are epigenetically determined. In all organisms, centromere location is specified by the assembly of unusual nucleosomes containing the histone H3 variant CENP-A in place of H3. However, an apparent paradox is that CENP-A in most organisms generally occurs on certain preferred sequences. The analyses presented focuses on the influence of DNA sequence on the selection of the locus where CENP-A chromatin are formed and whether there are any particular DNA features that promote CENP-ACnp1 assembly at centromeres in Schizosaccharomyces pombe. S. pombe provides an excellent model to study the structure and function of a complex eukaryotic centromere as it possesses epigenetically regulated centromeres that are structurally related to those of metazoa. Furthermore, plasmid-based circular minichromosomes provide a useful tool for studying these centromeres and the inactivation of minichromosome centromeres does not affect cell viability. The main aim of this study was to identify those features that enable centromeric sequences to assemble CENP-A chromatin. Manipulated circular minichromosomes were utilised to investigate the minimal central core sequence requirement for establishment of CENP-ACnp1 chromatin. These analyses showed that a minimal 2kb region from the central core of cen2 could form a functional centromere. A second aim was to analyse the DNA sequence requirements for centromere function on this minimal 2 kb region. To facilitate this, the endogenous central core region of endogenous cen2 was replaced with the central core region of cen1. This modified strain allows the structural and biological properties associated with plasmid borne central core 2 sequences to be analysed. Transcription of central domain sequences has been proposed to play a role in CENP-A establishment and/or maintenance. To explore the contribution of transcription potential promoters were mapped within the minimal 2 kb sequence and their regulatory elements investigated. Mutation of the minimal DNA element impedes its ability to assemble CENP-A chromatin. Therefore the primary DNA sequence of fission yeast centromeres is important for establishing functional centromeres and thus centromere location not entirely epigenetically regulated. It remains to be determined if the characteristics associated with theses sequences, and their mode of action, are conserved at other centromeres.

One form of protein regulation is accomplished by post-translational modification (PTM). In order to test the importance one type of PTM, methylation, in chromosome segregation, we inhibited protein methylation for brief periods in G2 using the general methylation inhibitor adenosine dialdehyde (AdOx). Inhibiting methylation solely in late G2 leads to mitotic defects. We observed that several methylated histone residues; H3K9me3, H4K20me3 and H4K20me1, are predominantly affected by AdOx in G2. We show both that the kinetochore proteins are not affected and that the mitotic checkpoint is intact. Further, we observed structural defects and chromosome misalignment in mitotic cells. These results indicate that methylation events during late G2 operate to maintain and ensure the structural integrity of pericentromeric heterochromatin prior to mitosis. These results suggest that pericentromeric heterochromatin is required for the proper sensing of kinetochore tension and inactivation of the mitotic checkpoint. / Experimental Oncology

Thesis (Ph. D.)--University of Missouri-Columbia, 2001. / Typescript. Vita. Includes bibliographical references (leaves 223-229). Also available on the Internet.

Thesis (Ph. D.)--University of Missouri-Columbia, 2001. / Typescript. Vita. Includes bibliographical references (leaves 223-229). Also available on the Internet.

Thesis (Ph. D.)--University of Washington, 2001. / Vita. Includes bibliographical references (leaves 85-103).

No description available.

Thesis (Ph.D.)--University of Melbourne, Dept. of Paediatrics, Faculty of Medicine, 2006. / Typescript. Includes bibliographical references (leaves 287-313).

Thesis advisor: Charles Hoffman / Faithful chromosome segregation is necessary for the successful completion of mitosis and meiosis. The centromere is the site of kinetochore and microtubule attachment during chromosome segregation, and it is critical that the centromere is properly formed and maintained. Many proteins contribute to centromere formation, and this process has been extensively studied during the mitotic cell cycle. However, the roles of the centromere and its associated proteins during meiosis and their contribution to the fidelity of chromosome segregation process are not as well understood. Here, I aim to elucidate a mechanism that may contribute to aneuploidy in gametes, which is a major contributing factor in human infertility. In this study, I investigate the role of Abp1, the most prominent member of the transposase-derived protein family homologous to mammalian CENP-B in the assembly of centromeric chromatin during meiosis in the fission yeast Schizosaccharomyces pombe. I reveal that in contrast to its known role as a major regulator of LTR retrotransposons during the mitotic and meiotic cell cycles, Abp1 has a specialized role at the centromere during meiosis. My results indicate that Abp1 displays dynamic localization to the centromeres during meiosis compared to the vegetative cell cycle. I show that loss of abp1 impairs pericentromeric heterochromatin and the localization of Cnp1, a CENP-A ortholog, to the centromere central cores during meiosis. Moreover, Abp1 appears to suppress formation of meiotic neocentromeres by restricting deposition of Cnp1 at certain heterochromatin loci. Loss of abp1 has a drastic effect on chromosome segregation, resulting in dramatic frequency of aneuploidy. Furthermore, the genome surveillance role for retrotransposons by Abp1 appears to encompass centromeres as the mere insertion of an LTR sequence within the centromere central cores further exacerbates incidence of meiotic aneuploidy in abp1 null cells. This study provides intriguing insights into factors controlling the assembly of centromeric chromatin and its impact on the fidelity of chromosome segregation process during meiosis with important implications for advancing our understanding of the evolutionary forces driving the evolution of eukaryotic centromeres. / Thesis (PhD) — Boston College, 2016. / Submitted to: Boston College. Graduate School of Arts and Sciences. / Discipline: Biology.