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。

基本信息

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

项目摘要

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 (PLD) 在多种类型细胞的信号转导中发挥着至关重要的作用，当细胞受到刺激时，它会被蛋白激酶 C α (PKCα) 激活。采用免疫组织学技术研究了肢端雀斑样黑色素瘤 (ALM) 和浅表扩散性黑色素瘤 (SSM) 原发性和转移性病变中 PLD1 和 PKCα 的表达。使用腺病毒介导的基因转移在人黑色素瘤细胞系 HM3KO 中检查了 PKCα 调节 PLD1 的机制，在 SSM 离线技术的原发性和转移性病变中，PLD1 和 DKCα 均强表达。这两种蛋白随着肿瘤的进展而急剧增加；在原发性ALM病灶的放射状生长期，PLD1和PKCα的表达几乎呈阴性，而在晚期则以与进展相关的方式同步增加。 ALM 病变，包括垂直生长期和转移性病变，免疫沉淀研究表明，在腺病毒介导的 PLD1 和 PKCα 或 PLD1 和PKCα 激酶阴性突变体揭示 PKCα 和 PKCα 激酶阴性突变体均与 PLD1 相关。在缺乏外部信号的情况下，腺病毒载体在黑色素瘤细胞中过度表达 PKCα 或 PKCα 激酶阴性突变体，导致基础 PLD 活性以病毒剂量依赖性方式增强。增加了 HM3KO 细胞的体外侵袭潜力。这些结果表明 PLD1 和 PKCα 的上调在 ALM 从径向生长期向垂直生长期的进展中发挥着重要作用。目前的结果还表明，PKCα 与 PLD1 相关，并以不依赖于蛋白质磷酸化的方式增强黑色素瘤细胞中的基础 PLD 活性，这有助于细胞的高侵袭潜力。