Morphological analysis of spherical extrusion induced in okadaic acid-treated cells by PEG-cholesterol

PEG-胆固醇诱导大田酸处理细胞球形挤压的形态学分析

• 批准号: 13670007
• 负责人: BABA Takeshi
• 金额: $ 2.62万
• 依托单位: University of Yamanashi, Faculty of Medicine
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Scientific Research (C)
• 财政年份: 2001
• 资助国家: 日本
• 起止时间: 2001 至 2002
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-13670007/
• 关键词: PEG-cholesterol; okadaic acid; actin microfilament; lipid raft; K562 cell; endocytosis; PEG-コレストロール; ダイナミン

Interconnection between surface microdomains and the actin cytoskeleton is vital to various cellular activities. We studied the responses of okadaic acid (OKA)-treated K562 leukemia cells to crosslinking of membrane microdomains. Although OKA alone induced clustering of surface-bound F-actin, addition of a biotinylated poly(ethylene glycol) derivative of cholesterol (bPEG-Chol) and subsequent binding of streptavidin (SA) further induced accumulation of the clusters, resulting in the formation of a spherical cell extrusion. This extrusion was also induced by direct crosslinking of a raft marker, CD59, and ganglioside GM1. In addition, we found that knockout of the gene encoding Fyn kinase inhibited formation of the spherical extrusion in murine T-cells. In bPEG-Chol/SA-treated cells, CD59, ganglioside GM1, and clathrin/AP-2 were all accumulated on the surface of the actin-rich extrusion, whereas dynamin and transferrin receptors were unaffected. Intermediate filaments, mitochondria, and other vesicles also accumulated. These results suggest that crosslinking of membrane domains exaggerates the linkage between actin and a defined set of membrane proteins in OKA-treated cells.

表面微区和肌动蛋白细胞骨架之间的相互连接对各种细胞活动至关重要。我们研究了冈田酸（OKA）处理的K562白血病细胞对膜微区交联的反应。尽管OKA单独诱导表面结合的F-肌动蛋白的聚集，但添加胆固醇的生物素化聚（乙二醇）衍生物（bPEG-Chol）和随后的链霉亲和素（SA）结合进一步诱导聚集，导致形成球形细胞挤出。这种挤出也是由筏标记物CD 59和神经节苷脂GM 1的直接交联引起的。此外，我们发现敲除编码Fyn激酶的基因抑制小鼠T细胞中球形挤出的形成。在bPEG-Chol/SA处理的细胞，CD 59，神经节苷脂GM 1，网格蛋白/AP-2都积累在表面上的肌动蛋白丰富的挤压，而发动蛋白和转铁蛋白受体不受影响。中间丝，线粒体和其他囊泡也积累。这些结果表明，交联的膜结构域夸大了肌动蛋白和一组定义的膜蛋白在OKA处理的细胞之间的联系。

项目成果

Yamaji-Hasegawa A, Makino A, Baba T, Senoh Y, Kimura-Suda H, Sato SB, Terada N, Ohno S, Kiyokawa E, Umeda M, Kobayashi T.: "Oligomerization and pore formation of a sphingomyelin-specific toxin, lysenin"J Biol Chem. (in press). (2003)

Yamaji-Hasekawa A、Makino A、Baba T、Senoh Y、Kimura-Suda H、Sato SB、Terada N、Ohno S、Kiyokawa E、Umeda M、Kobayashi T.：“鞘磷脂特异性毒素的寡聚化和孔形成，

Ueda H, Ueda K, Baba T, Ohno S: "d- and g-Sarcoglycan localization in the sarcoplasmic reticulum of skeletal muscle"J Histochem Cytochem. 49. 529-538 (2001)

Ueda H、Ueda K、Baba T、Ohno S：“骨骼肌肌浆网中的 d- 和 g-肌聚糖定位”J Histochem Cytochem。

Damke H, Binns DD, Ueda H, Schmid SL, Baba T: "Dynamin GTPase domain mutants block endocytic vesicle formation at morphologically distinct stages"Mol Biol Cell. 12. 2578-2589 (2001)

Damke H、Binns DD、Ueda H、Schmid SL、Baba T：“Dynamin GTPase 结构域突变体在形态不同的阶段阻断内吞囊泡形成”Mol Biol Cell。

Bata T, Udaka K, Terada N, Ueda H, Fujii Y, Ohno S, Sato SB: "Actin-rich spherical extrusion induced in okadaic acid-treated K562 cells by crosslinking of membrane microdomains"J Histochem Cytochem. 51(2). 245-252 (2003)

Bata T、Udaka K、Terada N、Ueda H、Fujii Y、Ohno S、Sato SB：“通过膜微域交联在冈田酸处理的 K562 细胞中诱导富含肌动蛋白的球形挤压”J Histochem Cytochem。

Makino A, Baba T, Fujimoto K, Iwamoto K, Yano Y, Terada N, Ohno S, Sato SB, Ohta A, Umeda M, Matsuzaki K, Kobayashi T: "Cinnamycin (Ro 09-0198) promotes cell binding and toxicity by inducing transbilayer lipid movement"J Biol Chem. 278(5). 3204-3209 (2003

Makino A、Baba T、Fujimoto K、Iwamoto K、Yano Y、Terada N、Ohno S、Sato SB、Ohta A、Umeda M、Matsuzaki K、Kobayashi T：“肉桂霉素 (Ro 09-0198) 通过以下方式促进细胞结合和毒性：