Regulation of cross-linking of actin filament by IQGAP1, a target for Cdc42
Regulation of cross-linking of actin filament by IQGAP1, a target for Cdc42
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DOI：
10.1074/jbc.272.47.29579
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发表时间：
1997-11-21
影响因子：
4.8
通讯作者：
Kaibuchi, K
中科院分区：
文献类型：
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作者：
Fukata, M;Kuroda, S;Kaibuchi, K
我们先前已经表明，IQGAP 1，一种最近鉴定的Cdc 42和Rad小GTP酶的靶标，在由胰岛素和Rac 1（val 12）诱导的膜皱褶区域显示出与皮质肌动蛋白细胞骨架相似的分布（黑田，S.，Yakata，M，，小林，K.，Nakafuku，M.，野村，N.，Iwamatsu，A.，和Kaibuchi，K.（1996）J.Biol.Chem.271，28363-23367）。在这里，我们确定了IQGAP 1相互作用的分子量为43 kDa（p43）从牛脑胞质溶胶，使用谷胱甘肽S-转移酶（GST）-IQGAP 1亲和柱层析。蛋白质的氨基酸序列分析显示，p43与β-和γ-肌动蛋白相同。IQGAP 1与丝状肌动蛋白（F-actin）共沉淀。IQGAP 1的氨基端结构域（氨基酸1-216）负责与F-肌动蛋白的相互作用。落球粘度测定显示IQGAP 1交联了F-肌动蛋白。这种IQGAP 1活性被鸟苷5 ′-（3-O-硫代）三磷酸（GTP γ S）GST-Cdc 42进一步增强，但不被GDP-GST-Cdc 42增强。IQGAP 1的凝胶过滤分析显示IQGAP 1表现为寡聚体，并且GTP γ S-GST-Cdc 42而不是GDP GST-Cdc 42增强IQGAP 1的寡聚化。这些结果有力地表明，IQGAP 1，作为下游的Cdc 42，可以交联肌动蛋白丝通过其寡聚化。
We have previously shown that IQGAP1, a recently identified target for Cdc42 and Rad small GTPases, showed a distribution similar to that of cortical actin cytoskeleton at the membrane ruffling area induced by insulin and Rac1(val12) (Kuroda, S., Fukata, M,, Kobayashi, K., Nakafuku, M., Nomura, N., Iwamatsu, A., and Kaibuchi, K. (1996) J. Biol. Chem. 271, 28363-23367). Here we identified an IQGAP1-interacting molecule with molecular mass of 43 kDa (p43) from bovine brain cytosol, using glutathione S-transferase (GST)-IQGAP1 affinity column chromatography. The amino acid sequencing of the protein revealed that p43 was identical to beta- and gamma-actin. IQGAP1 was cosedimentated with filamentous actin (F-actin). The amino-terminal domain (amino acids 1-216) of IQGAP1 was responsible for the interaction with F-actin. Falling ball viscometry assay revealed that IQGAP1 cross-linked the F-actin. This IQGAP1 activity was further enhanced by guanosine 5'-(3-O-thio)triphosphate (GTP gamma S) GST-Cdc42 but not by GDP-GST-Cdc42. The gel filtration analysis of IQGAP1 revealed that IQGAP1 appeared as oligomers and that GTP gamma S-GST-Cdc42 but not GDP GST-Cdc42 enhanced the oligomerization of IQGAP1. These results strongly suggest that IQGAP1, acting downstream of Cdc42, can cross-link the actin filament through its oligomerization.