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Title: Trafficking and assembly of band 3 based multiprotein complexes during erythropoiesis
Author: Satchwell, Timothy James
ISNI:       0000 0004 2725 5177
Awarding Body: University of Bristol
Current Institution: University of Bristol
Date of Award: 2011
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The work presented in this thesis has developed an existing in vitro culture system used for expansion of cord blood progenitors to one that can be applied to expand significant numbers of erythroblasts from peripheral blood mononuclear cells isolated from healthy donor blood samples or patients with haemolytic anaemia. These erythroblasts can be further differentiated to enucleated reticulocytes synchronously producing homogeneous populations of cells at each of the morphologically defined stages of erythroid differentiation in quantities amenable to biochemical experimentation. Expression profiles of proteins that constitute the band 3 based multiprotein complex were monitored throughout-erythropoiesis by a variety of methods and these studies were extended to investigate the spatial and temporal establishment of critical interactions required for multiprotein complex assembly during differentiation of erythroblasts to reticulocytes. Protein 4.2, 'a key component of this complex was found to be expressed early during terminal erythroid differentiation in basophilic erythroblasts where it interacts with band 3 in an intracellular compartment prior to delivery to the plasma membrane. Rh and its associated glycoprotehRhAG which form the core of the band 3 associated Rh complex were also found to interact at this stage at the plasma membrane simultaneous to the onset of Rh dependency on RhAG. Expansion and differentiation of erythroblasts derived from a protein 4.2 null hereditary spherocytosis patient was conducted for the first time. This demonstrated that the hallmarks of protein 4.2 absence observed in erythrocytes begin to manifest within 48 hours of differentiation concomitant with the onset of initial complex assembly in basophilic erythroblasts and within a window of major membrane protein synthesis and delivery. Initial culture of erythroblasts from CDAII patients suggests that the onset of a glvcosvlation defect also occurs from the onset of terminal differentiation whilst additional characteristic phenotypes of this disease develop later.
Supervisor: Not available Sponsor: Not available
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral
EThOS ID:  DOI: Not available