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Degree

Doctor of Philosophy, Case Western Reserve University, Biology, 1990.

Abstract

This study has used several systems to characterize (i) the transport of membrane proteins to and from the cell surface, and (ii) the maintenance and structural organization of the Golgi complex. The transport of membrane proteins was studied using the regulated surface expression of the C3b/C4b receptor, CR1, on human neutrophils as a model. We developed a new method for quantitative immunofluorescence of intracellular antigens, and used this approach to document upregulation of cell surface CR1 and to identify ligand-independent degradation of CR1. This is the first example of acute ligand-independent receptor degradation. Light and electron microscopic immunocytochemistry resulted in the description of a previously unrecognized granule as the site of intracellular CR1 storage in neutrophils, and the identification of multivesicular bodies as a possible site of CR1 degradation. The Golgi complex is composed of multiple cisternae assembled into stacks which form a single interconnected organelle. Thus, the Golgi complex was investigated to study the maintenance of organelle structural organization. The first approach was biochemical and involved identification of Golgi-associated cytoskeletal material potentially important in cisternal stacking or maintenance of cisternal shape. Five proteins were isolated, but the data could not conclusively demonstrate the existence or function of a Golgi-associated cytoskeleton. Nonetheless, immunization of mice with these proteins and preparation of monoclonal antibodies resulted in the production of an antibody which recognizes epitopes on intermediate filaments, actin stress fibers, and previously unidentified cytoplasmic foci. It is possible that this antibody recognizes a structure involved in integration of these separate components of the cytoskeleton into a unified system. The association of Golgi stacks with microtubules and the organization of these stacks into a single Golgi complex was also examined. The data demonstrate that reversible fragmentation of the coherent juxtanuclear Golgi follows microtubule depolymerization, apparently as a result of membrane traffic and other energy-dependent processes. Golgi organization and microtubule status were clearly dissociable, thereby proving that organization of individual small stacks into a single interconnected Golgi is an active process not simply determined by microtubule binding.

Keywords

Secretory organelles cytoskeleton Organization interdependence

Advisor

Alan M. Tartakoff

Pages

183p.

Document number: case1054668919
Permalink: http://rave.ohiolink.edu/etdc/view?acc_num=case1054668919

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