Cyclic phosphatidic acid inhibits RhoA-mediated autophosphorylation of FAK at Tyr-397 and subsequent tumor-cell invasion

被引:4
作者
Mukai, M
Iwasaki, T
Tatsuta, M
Togawa, A
Nakamura, H
Murakami-Murofushi, K
Kobayashi, S
Imamura, F
Inoue, M
机构
[1] Osaka Med Ctr Canc & Cardiovasc Dis, Dept Tumor Biochem, Higashinari Ku, Osaka 5378511, Japan
[2] Osaka Med Ctr Canc & Cardiovasc Dis, Dept Gastrointestinal Oncol, Osaka 5378511, Japan
[3] Osaka Med Ctr Canc & Cardiovasc Dis, Dept Pulm Oncol, Osaka 5378511, Japan
[4] Osaka Prefectural Habikino Hosp, Dept Resp Surg, Osaka 5838588, Japan
[5] Ochanomizu Univ, Fac Sci, Dept Biol, Tokyo 1128610, Japan
[6] Sci Univ Tokyo, Fac Pharmaceut Sci, Med Chem Lab, Tokyo 1620826, Japan
关键词
cyclic phosphatidic acid; lysophosphatidic acid; Rho; focal adhesion kinase; tumor invasion;
D O I
暂无
中图分类号
R73 [肿瘤学];
学科分类号
100214 ;
摘要
We demonstrated previously that rat ascites hepatoma MM1 cells require both lysophosphatidic acid (LPA) and fibronectin (FN) for phagokinetic motility and transcellular migration and that these events are regulated through the RhoA-ROCK pathway and tyrosine phosphorylation of proteins including focal adhesion kinase (FAK). Moreover, we reported that palmitoyl-cyclic phosphatidic acid (Pal-cPA), a structural analogue of LPA, inhibits LPA-induced migration of MMI cells and experimental metastasis of B16 murine melanoma cells. However, the molecular mechanisms of action of Pal-cPA remains to be clarified. To examine this, total cellular lysates after stimulation with LPA or FN were subjected to time-course immunoblot analysis with anti-phophotyrosine and anti-pY397-FAK antibodies. Tyrosine-phosphorylation of FAK especially at Tyr-397 was obviously persistent after stimulation with LPA + FN compared to after stimulation with LPA alone. This persistent phosphorylation was necessary for MMI cell migration and inhibited by Pal-cPA as by C3 exoenzyme Rho inhibitor. RhoA activity (GTP-bound RhoA) was also measured by the pull down assay using the Rho binding domain of Rhotekin. LPA-induced RhoA-activation of MMI cells was completely inhibited by Pal-cPA. Moreover, we demonstrated that autophosphorylation of FAK at Tyr-397, downstream of RhoA, contributed to formation of focal adhesions and was critical in LPA-induced MMI cell migration by developing autophosphorylation-defficient (Y397F) FAK-transfectants. Collectively, Pal-cPA hampered LPA-induced morphological changes and transcellular migration of MMI cells through downregulating active RhoA and inhibiting its downstream events including autophosphorylation of FAK. Pal-cPA also inhibited endogenous (LPA-independent) activation of RhoA in human fibrosarcoma HT-1080 cells. Pal-cPA may potentially provide a new therapy for the treatment of cancer invasion and metastasis.
引用
收藏
页码:1247 / 1256
页数:10
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