学位论文详细信息
MOLECULAR MECHANISMS REGULATING COPPER BALANCE IN HUMAN CELLS
copper;ATP7A;ATP7B;Ctr1;ANKRD9;Physiology
Hasan, Nesrin MLutsenko, Svetlana ;
Johns Hopkins University
关键词: copper;    ATP7A;    ATP7B;    Ctr1;    ANKRD9;    Physiology;   
Others  :  https://jscholarship.library.jhu.edu/bitstream/handle/1774.2/60309/HASAN-DISSERTATION-2014.pdf?sequence=1&isAllowed=y
瑞士|英语
来源: JOHNS HOPKINS DSpace Repository
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【 摘 要 】

Precise copper balance is essential for normal growth, differentiation, and function of human cells. Loss of copper homeostasis is associated with heart hypertrophy, liver failure, neuronal de-myelination and other pathologies. The copper-transporting ATPases ATP7A and ATP7B maintain cellular copper homeostasis. In response to copper elevation, they traffic from the trans-Golgi network (TGN) to vesicles where they sequester excess copper for further export. The mechanisms regulating activity and trafficking of ATP7A/7B are not well understood. Our studies focused on determining the role of kinase-mediated phosphorylation in copper induced trafficking of ATP7B, and identifying and characterizing novel regulators of ATP7A. We have shown that Ser-340/341 region of ATP7B plays an important role in interactions between the N-terminus and the nucleotide-binding domain and that mutations in these residues result in vesicular localization of the protein independent of the intracellular copper levels. We have determined that structural changes that alter the inter-domain interactions initiate exit of ATP7B from the TGN and that the role of copper-induced kinase-mediated hyperphosphorylation might be to maintain an open interface between the domains of ATP7B. In a separate study, seven proteins were identified, which upon knockdown result in increased intracellular copper levels. We performed an initial characterization of the knock-downs and obtained intriguing results indicating that these proteins regulate ATP7A protein levels, post-translational modifications, and copper-dependent trafficking.None of these proteins has been previously linked to copper homeostasis. Our studies have pointed to novel kinase inhibitors (IBTK and CAMK2N2), a novel relationship between copper transport and organic anion/bile acid transport (ABCC3), a likely mechanism of ATP7A regulation through changes in glycosylation levels, and identified an adaptor protein (ankyrin repeat domain protein 9 (ANKRD9)) that might be involved in stabilizing the Golgi structure. ANKRD9 depletion led to increase of intracellular copper levels, Golgi fragmentation, and defects in protein glycosylation. Additionally, the mRNA and protein levels of the human copper uptake protein Ctr1 were increased significantly despite high copper levels in the cells. We are facing challenging questions related to Golgi fragmentation and copper homeostasis. Further characterization of the above-mentioned novel regulatory modes will greatly expand our understanding of copper metabolism in human cells.

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