跳到主要內容

臺灣博碩士論文加值系統

(3.236.23.193) 您好!臺灣時間:2021/07/26 07:50
字體大小: 字級放大   字級縮小   預設字形  
回查詢結果 :::

詳目顯示

我願授權國圖
: 
twitterline
研究生:林芳如
研究生(外文):Fan-Ru Lin
論文名稱:NuMA蛋白的cdc2kinase磷酸化位置之突變影響有絲分裂細胞週期之進行
論文名稱(外文):Mutations of the cdc2 kinase phosphorylation sites of NuMA alters cell cycle progression in mitosis.
指導教授:葉寧馨
指導教授(外文):Ning-Hsing Yeh
學位類別:碩士
校院名稱:國立陽明大學
系所名稱:微生物及免疫學研究所
學門:生命科學學門
學類:微生物學類
論文種類:學術論文
論文出版年:2004
畢業學年度:92
語文別:中文
論文頁數:85
中文關鍵詞:NuMA蛋白cdc2磷酸化激酶有絲分裂
外文關鍵詞:NuMAcdc2 kinasemitosis
相關次數:
  • 被引用被引用:0
  • 點閱點閱:86
  • 評分評分:
  • 下載下載:0
  • 收藏至我的研究室書目清單書目收藏:0
uMA (Nuclear Mitotic Apparatus Protein)在細胞中的分布位置以及所扮演的角色會隨著細胞週期改變而有所不同。在間期細胞中,NuMA蛋白分布於整個細胞核除核仁以外的區域,被認為是核基質的一員或為核基質之結合蛋白,主要參與在維持細胞核結構之構造組成;而當細胞進入有絲分裂期時,NuMA蛋白會被磷酸化,並離開濃縮之染色質快速聚集至紡綞體頂端區域,主要參與在幫助及維持紡綞體之完整性。因為NuMA蛋白在有絲分裂時期被磷酸化與其在有絲分裂時期的分布變化有時間上之一致性,而NuMA蛋白的C-端有四個預測的cdc2 kinase磷酸化位置,並且cdc2 kinase的活化會促使有絲分裂的進行,因此想知道在有絲分裂時期是否透過cdc2 kinase將NuMA蛋白磷酸化,而導致NuMA蛋白分布的改變。分別建構逐漸增加cdc2 kinase磷酸化位置突變數於NuMA cDNA上。發現當NuMA蛋白上的cdc2 kinase磷酸化位置有兩個以上被突變,會導致此突變蛋白在有絲分裂時與紡綞體結合的能力下降,並且隨著突變的cdc2 kinase磷酸化位置越多,此突變蛋白失去與紡綞體結合特性就越明顯。並且觀察到當細胞表現此突變蛋白會影響細胞週期的進行,導致prometaphase在有絲分裂期細胞群中比例明顯增加,顯示著NuMA在有絲分裂被cdc2 kinase磷酸化可能扮演著一個新的調控機制,調控細胞從prometaphase進行至metaphase。
另外,NuMA蛋白的C-端已經被報導知道帶有許多的功能性區域,我建構了幾個含有尾端之截短NuMA。發現只含有C-端球狀結構的NuMA截短蛋白,並不會聚集至紡綞體兩端;而含有C-端球狀蛋白前端以及其鄰近部分coiled-coil區域之NuMA截短蛋白則是有時會聚集至紡綞體,有時則散佈於細胞中;而只缺少與C-端球狀結構相鄰之部分coiled-coils區域的NuMA截短蛋白則完全散佈於細胞中。除了這些截短蛋白在細胞中有不正常分布之外,並未觀察到對細胞有任何不正常的影響。結果顯示C-端球狀結構並沒有具有使NuMA蛋白與紡綞體頂端相結合的能力,而是與C-端球狀結構相鄰的coiled-coils區域對於NuMA蛋白聚集至紡綞體兩端的能力是需要的。
NuMA (Nuclear Mitotic Apparatus Protein) distributes in the whole nucleus except the nucleolus in interphase, but translocates to the spindle poles in mitosis. NuMA is regarded as a component of nuclear matrix, and is responsible for nuclear structural maintenance in interphase. In mitosis, NuMA is involved in spindle organization and stabilization. There are four predicted cdc2 kinase phosphorylation sites at the C-terminal of NuMA. It is known that activation of cdc2 kinase promotes the onset of mitosis. The question whether the cdc2-mediated phosphorylation of NuMA correlates with its redistribution in mitosis was tested. The full-length NuMA cDNAs with increased number of mutations at cdc2 phosphorylation sites were constructed. These NuMA mutants with more than two sites to be mutated reduced their abilities to associate with spindle poles. Decrease of the spindle-association became more significant if more than three cdc2 kinase phosphorylation sites were mutated. Besides, these mutant NuMA proteins resulted in accumulation of prometaphase population. Such results indicate that cdc2-mediated phosphorylation of NuMA might serve as a new mitotic checkpoint for cells passing from prometaphase to metaphase.
It is known that many functional domains of NuMA are located at its C-terminus. Several deletion clones containing parts of the NuMA C-terminal end were constructed for experiments to observe phenotypes of cells expressing these NuMA deletion mutants. The globular tail domain alone did not show accumulation in spindle poles. Deletion mutants of NuMA missing only part of the coiled-coils adjacent to the globular tail dispersed in the whole cell without translocation to spindle poles. Besides the abnormal distributions of these deletion NuMA, such tansfectants did not show any abnormal phenotypes. It indicates that the C-terminal globular end of NuMA alone does not have the ability to associate with spindle poles. However, the adjacent coiled-coils region is necessary for spindle association.
Compton, D. A. and D. W. Cleveland, 1993. "NuMA is required for the proper
completion of mitosis." J Cell Biol 120(4): 947-57.
Compton, D. A. and C. Luo, 1995. "Mutation of the predicted p34cdc2
phosphorylation sites in NuMA impair the assembly of the mitotic spindle and block
mitosis." J Cell Sci 108 ( Pt 2): 621-33.
Compton, D. A., I. Szilak, et al. 1992. "Primary structure of NuMA, an intranuclear
protein that defines a novel pathway for segregation of proteins at mitosis." J Cell
Biol 116(6): 1395-408.
Christiana, M., Caroline, R., Stephanie, M. T., Solange, M., Sylvie, K., Jerome,
G.,Paul, R. A., Annick, B., Bruno, G., Jean, P. K., Robert L.M., 2003. "The
mammalian passenger protein TD-60 is an RCC1 family member with an essential
role in prometaphase to metaphase progression." Developmental Cell 5: 295-307.
Du, Q., L. Taylor, Compton, D. A., Macara, I. G., 2002. "LGN blocks the ability of
NuMA to bind and stabilize microtubules. A mechanism for mitotic spindle assembly
regulation." Curr Biol 12(22): 1928-33.
D. W. Cleveland, Yinghui M., K. F. Sullivan, 2003. "Centromeres and kinetochores:
from epigenetics to mitotic checkpoint signaling. "Cell 112: 407-421.
Gaglio, T., A. Saredi, Compton D. A., 1995. "NuMA is required for the organization
of microtubules into aster-like mitotic arrays." J Cell Biol 131(3): 693-708.
Gehmlich, K., L. Haren, Merdes, A., 2004. "Cyclin B degradation leads to NuMA
release from dynein/dynactin and from spindle poles." EMBO Rep 5(1): 97-103.
Harborth, J., J. Wang, Gueth-Hallonet, C., Weber, K., Osborn, M., 1999. "Self
assembly of NuMA: multiarm oligomers as structural units of a nuclear lattice." Embo
J 18(6): 1689-700.
Haren, L. and A. Merdes, 2002. "Direct binding of NuMA to tubulin is mediated by
a novel sequence motif in the tail domain that bundles and stabilizes microtubules." J
Cell Sci 115(Pt 9): 1815-24.
M. A. Hoyt, 2001. "A new view of the spindle checkpoint." J. Cell Biol. 154:
909-911.
Hsu, H. L. and N. H. Yeh, 1996. "Dynamic changes of NuMA during the cell cycle
and possible appearance of a truncated form of NuMA during apoptosis." J Cell Sci
109 ( Pt 2): 277-88.
J. V. Shah, D.W. Cleveland, 2000. "Waiting for anaphase: Mad2 and the spindle
assembly checkpoint." Cell 103: 997-1000.
Kallajoki, M., K. Weber, Osborn, M., 1991. "A 210 kDa nuclear matrix protein is a
functional part of the mitotic spindle; a microinjection study using SPN monoclonal
antibodies." Embo J 10(11): 3351-62.
Kallajoki, M., K. Weber, Osborn, M., 1992. "Ability to organize microtubules in
taxol-treated mitotic PtK2 cells goes with the SPN antigen and not with the
centrosome." J Cell Sci 102 ( Pt 1): 91-102.
Lydersen, B. K. and D. E. Pettijohn, 1980. "Human-specific nuclear protein that
associates with the polar region of the mitotic apparatus: distribution in a
human/hamster hybrid cell." Cell 22(2 Pt 2): 489-99.
Merdes, A., R. Heald, Samejima, K., Earnshow, W., C. Cleveland, D. W., 2000.
"Formation of spindle poles by dynein/dynactin-dependent transport of NuMA." J
Cell Biol 149(4): 851-62.
Merdes, A., K. Ramyar, Vechio, J. D., Cleveland, D. W., 1996. "A complex of
NuMA and cytoplasmic dynein is essential for mitotic spindle assembly." Cell 87(3):
447-58.
R. Bharadwaj and H. Yu, 2004. "The spindle checkpoint, aneuploidy, and cancer."
Oncogene 23: 2016-2027.
Stewart, M., 1990. "Intermediate filaments: structure, assembly and molecular
interaction." Curr. Opin, Cell Biol. 2: 91-100
Tajao Ono, Ana Losada, Michiko Hirano, Michael P. Myers, Andrew F. Neuwald,
Tatsuya Hirano, 2003. "Differential contributions of condesin I and condesin II to
mitotic chromosome architecture in vertebrate cells." Cell 115: 109-121
Tang, T. K., C. J. Tang, Chao, Y. J., C. W., 1994. "Nuclear mitotic apparatus protein
(NuMA): spindle association, nuclear targeting and differential subcellular
localization of various NuMA isoforms." J Cell Sci 107 ( Pt 6): 1389-402.
Tang, T. K., C. J. Tang, Chen, Y. J., Wu, C. W., 1993. "Nuclear proteins of the
bovine esophageal epithelium. II. The NuMA gene gives rise to multiple mRNAs and
gene products reactive with monoclonal antibody W1." J Cell Sci 104 ( Pt 2): 249-60.
Tousson, A., C. Zeng, Brinkly, B. R., Valdivia, M. M., 1991. "Centrophilin: a novel
mitotic spindle protein involved in microtubule nucleation." J Cell Biol 112(3):
427-40.
Yang, C. H., E. J. Lambie, Snyder, M., 1992. "NuMA: an unusually long coiled-coil
related protein in the mammalian nucleus." J Cell Biol 116(6): 1303-17.
Zeng, C., He, D., Brinkley, B. R., 1994. "Localization of NuMA protein isoforms in
the nuclear matrix of mammalian cells." Cell Motil Cytoskeleton 29(2): 167-76.
QRCODE
 
 
 
 
 
                                                                                                                                                                                                                                                                                                                                                                                                               
第一頁 上一頁 下一頁 最後一頁 top