Study of the role of protein kinase C-phosphoipase D signaling pathway in pigment cells.

蛋白激酶C-磷酸酶D信号通路在色素细胞中的作用研究。

• 批准号: 13670886
• 负责人: OKA Masahiro
• 金额: $ 2.3万
• 依托单位: Kobe University
• 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-13670886/
• 关键词: malignant melanoma; protein kinase C; phosphorylation; protein-protein interaction; invasion; 色素細胞; Cキナーゼ; ホスホリパーゼD

It is well known that phospholipase D (PLD) plays a crucial role in the signal transduction of many types of cells, and is activated by protein kinase C α (PKCα) when cells are stimulated. To elucidate the role of PLD in melanoma, the expression of PLD1 and PKCα in primary and metastatic lesions of acral lentiginous melanoma (ALM) and superficial spreading melanoma (SSM) was investigated using immunohistological techniques. In addition, the mechanism of regulation of PLD1 by PKCα was examined in a human melanoma cell line HM3KO using an adenovirus-mediated gene transfer technique. Both PLD1 and DKCα were strongly expressed in primary and metastatic lesions of SSM. Conversely, in ALM lesions, the expression of these two proteins increased dramatically with tumor progression ; the expression of both PLD1 and PKCα was almost negative in the radial growth phase of primary ALM lesions, and increased synchronously in a progression-related manner in advanced ALM esions, including vertical gro … More wth phase and metastatic lesions. Immunoprecipitation study showed that PLD1 and PKCα are associated physiologically in resting melanoma cells. Further immunoprecipitation study using HM3KO cells after adenovirus-mediated simultaneous overexpression of PLD1 and PKCα, or PLD1 and the kinase-negative mutant of PKCα revealed that both PKCα and the kinase-negative mutant of PKCα are associated with PLD1 in melanoma cells in the absence of an extemal signal. Overexpression of PKCα or the kinase-negative mutant of PKCα in melanoma cells by the adenovirus vectors resulted in the enhancement of basal PLD activity in a viral dose-dependent manner. Furthermore, enhanced basal PLD activity increased the in vitro invasive potential of HM3KO cells. These results suggest that upregulation of PLD1 and PKCα plays a rote in the progression of ALM from the radial growth phase to the vertical growth phase. The present results also suggest that PKCα associates with PLD1 and enhances basal PLD activity in a protein phosphorylation-independent manner in melanoma cells, which contributes to the cell's high invasive potential. Less

众所周知，磷脂酶D在多种细胞的信号转导中起重要作用，在细胞受到刺激时被蛋白激酶Cα(PKCα)激活。为探讨磷脂酶D在黑色素瘤中的作用，采用免疫组织化学方法检测了磷脂酶D 1和蛋白激酶Cα在肢端狼疮性黑色素瘤和浅表性播散性黑色素瘤原发灶和转移灶中的表达。此外，用腺病毒介导的基因转移技术在人黑色素瘤细胞系HM3KO中研究了PKCα对pld1基因调控的机制。α在SSM原发灶和转移灶中均呈强阳性表达。相反，在ALM皮损中，这两种蛋白的表达随着肿瘤的进展而显著增加；在原发ALM皮损的放射状生长期，pld1和pkcα的表达几乎为阴性，而在晚期ALM中，包括垂直型…，两者的表达与进展相关。分期和转移灶较多。免疫沉淀研究表明，在静息的黑色素瘤细胞中，pld1和pKCα在生理上是相关的。进一步的免疫沉淀研究表明，在没有外界信号的情况下，在黑色素瘤细胞中，pLd1和pkCα同时过表达后，hm3ko细胞与pLd1和pKCα的激酶阴性突变体pKCα和pLd1相关。腺病毒载体在黑色素瘤细胞中过表达PKCα或PKCα的突变株后，其基础PLD活性呈病毒剂量依赖性增强。此外，基础PLD活性的增强增加了HM3KO细胞的体外侵袭能力。这些结果表明，在ALM从径向生长期到垂直生长期的过程中，pld1和pKCα的上调起着重要作用。目前的结果还表明，在黑色素瘤细胞中，PKCα与pld1结合并以不依赖于蛋白质磷酸化的方式增强基础pld活性，这有助于黑色素瘤细胞的高侵袭性。较少

项目成果

Oka, M.: "Dual regulation of phospholipase D1 by protein kinase Cα in vivo"Biochem. Biophys. Res. Commun.. 294・5. 1109-1113 (2001)

Oka, M.：“体内蛋白激酶 Cα 的双重调节”Biochem. Commun. 1109-1113 (2001)。

Oka, M.: "Dual regulation of phospholipase D1 by protein kinase Cα in vivo"Biochem.Biophys.Res.Commun.. 294. 1109-1113 (2002)

Oka, M.：“体内蛋白激酶 Cα 对磷脂酶 D1 的双重调节”Biochem.Biophys.Res.Commun. 294. 1109-1113 (2002)

Nagai, H.: "Gene transfer of secreted-type modified interleukin-18 gene to B16F10 melanoma cells suppresses in vivo tumor growth through inhibition of tumor vessel formation"J. Invest. Dermatol.. 119・3. 541-548 (2002)

Nagai, H.：“将分泌型修饰的白细胞介素 18 基因转移至 B16F10 黑色素瘤细胞，通过抑制肿瘤血管形成来抑制体内肿瘤生长”J. Invest. 119・3 (2002)。

Mizuho Fukunaga et al.: "UV-Induced Tyrosine Phosphorylation of PKCS and Promstion of Apoptosis in the HaCaT Cell Line"Biochemical And Biophysical Research Communications. 289・2. 573-579 (2001)

Mizuho Fukunaga 等人：“HaCaT 细胞系中 PKCS 的紫外线诱导酪氨酸磷酸化和细胞凋亡的促进”生物化学和生物物理研究通讯 289・2（2001）。

Nagai, H.: "Gene transfer of secreted-type modified interleukin-18 gene to B16F10 melanoma cells suppresses in vivo tumor growth through inhibition of tumor vessel formation"J.Invest Dermatol.. 119. 541-548 (2002)

Nagai, H.：“将分泌型修饰的白细胞介素 18 基因转移至 B16F10 黑色素瘤细胞，通过抑制肿瘤血管形成来抑制体内肿瘤生长”J.Invest Dermatol.. 119. 541-548 (2002)