Analysis of the molecular structure of centromeres of Chinese hamster and human chromosomes.
thesisposted on 19.11.2015, 08:53 by Katalin Anna. Holland
Monoclonal anti-CHO chromosome scaffold, and polyclonal anti-centromere autoimmune antibodies were employed to investigate the molecular organisation of the centromeres of Chinese hamster and human chromosomes. The presence of a novel ring-like structure was revealed, which was conserved amongst the metaphase centromeres of two rodents, namely Chinese hamster and mouse, and in humans. In metaphase the antigen had a molecular weight of 170kD, this polypeptide may be processed from, or the stable degradation product of, a 200kD interphase precursor full-length polypeptide. The antigen could not be detected in interphase centromeres, but the ring-like structure was detected in chromosome scaffold preparations, indicating that the ring antigen resisted the 2M NaCl extraction, and hence, tightly bound the DNA of metaphase chromosomes. The possible relationship of the centromere antigen with another high molecular weight and major chromosome scaffold protein, DNA topoisomerase II, was also explored. The centromere antigen was also localised apparently to the interdigitating microtubules of the midzone, in anaphase; seemingly to the contractile ring at cytokinesis, and also to the interphase centrioles. The antigen was shown to be distinct from the microtubule-associated proteins and had non-identity with another high molecular weight protein component of the contractile ring, namely myosin. The dramatic relocation of the antigen, however, at the metaphase/anaphase transition suggests that it falls into a class of recently identified dual chromosomal and cytoplasmic components, termed the "passenger proteins". The ring antigen is postulated to be involved in holding the sister chromatids together by a strong interaction with the centromeric DNA through prometaphase to the metaphase-anaphase transition. It is suggested that the ring antigen is disengaged from the centromere prior to anaphase, and may be involved in the regulation of the metaphase-anaphase transition.