Title page for etd-0910107-105059


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URN etd-0910107-105059
Author Ching-chih Lin
Author's Email Address irvin1971@hotmail.com
Statistics This thesis had been viewed 5586 times. Download 15 times.
Department Biological Sciences
Year 2006
Semester 2
Degree Ph.D.
Type of Document
Language English
Title Multiple tasks of Glycogen synthase kinase-3beta (GSK-3β ) and its partners
Date of Defense 2007-07-30
Page Count 111
Keyword
  • neurite
  • Glycogen synthase kinase-3
  • neuroblastoma
  • centrosomal protein
  • GSKIP
  • gamma-tubulin docking sites
  • Abstract Glycogen synthase kinase-3 (GSK-3) is a serine/threonine protein kinase which plays a key role in several signaling pathways and its homologues have been identified in most eukaryotes. Since GSK3βis an essential protein kinase that regulates numerous functions within the cell, an effort to survey possible GSK3β- interacting proteins from a human testis cDNA library using the yeast two-hybrid system is made. Two interesting candidates are chosen to characterize their functions in this study. One is a centrosomal protein, hNinein, and the other is a novel inhibitor of GSK3β, designated as GSKIP (GSK3β interaction protein).
    In the first part of the present thesis we describe the identification of four diverse CCII-termini of human hNinein isoforms, including a novel isoform 6, by differential expression in a tissue-specific manner. In a kinase assay, the CCII region of hNinein isoforms provides a differential phosphorylation site by GSK3β. In addition, either N-terminal or CCIIZ domain disruption may cause hNinein conformational change which recruits γ-tubulin to centrosomal or non-centrosomal hNinein-containing sites. Further, depletion of all hNinein isoforms caused a significant decrease in the γ-tubulin signal in the centrosome. In domain swapping, it clearly shows that the CCIIX-CCIIY region provides docking sites for γ-tubulin. Moreover, nucleation of microtubules from the centrosome is significantly affected by the overexpression of either the full-length hNinein or CCIIX-CCIIY region. Taken together, these results show that the centrosomal targeting signals of hNinein have a role not only in regulating hNinein conformation, resulting in localization change, but also provide docking sites to recruit γ-tubulin at centrosomal and non-centrosomal sites.
    In the second part of the thesis we describe another candidate, GSK3βinteraction protein (GSKIP), to characterize its functions in neuron differentiation. We use human neuroblastoma SH-SY5Y cells as a model of neuronal cell differentiation. When overexpression of GSKIP prevents neurite outgrowth from RA-mediated differentiation, this result is similar to the presence of LiCl or SB415286, an inhibitor of GSK3β. Further, GSKIP regulates the activity of GSK3β through protein-protein interactions rather than post-modulation and GSKIP may affect GSK3β on neurite outgrowth via inhibiting the specific phosphorylation site of tau. In addition to inhibition of neurite outgrowth, GSKIP overexpressed in SH-SY5Y cells also promotes cell cycle progression by analyzing cell proliferation with cell growth and MTT assay. Furthermore, GSKIP raises the level of β-catenin and cyclin D1 through inhibition of GSK3β activity in RA-mediated differentiation SH-SY5Y cells. Taken together, the data suggest that GSKIP, a dual functional molecule, is able to inhibit neurite outgrowth and promote cell proliferation via negative regulation of GSK3β activity in RA-mediated differentiation of SH-SY5Y cells.
    Advisory Committee
  • Chung-Lung Cho - chair
  • Pei-Jung Lu - co-chair
  • De-Ching Chang - co-chair
  • Wen-Tsan Chang - co-chair
  • Bei-Chang Yang - co-chair
  • Yi-Ren Hong - advisor
  • Ching-Mei Hsu - advisor
  • Files
  • etd-0910107-105059.pdf
  • indicate in-campus access in a year and off_campus not accessible
    Date of Submission 2007-09-10

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