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

磷脂酶D（phospholipase D，PLD）在多种细胞的信号转导中起重要作用，当细胞受到刺激时，PLD被蛋白激酶C α（protein kinase C α，PKCα）激活。为探讨PLD在黑色素瘤中的作用，采用免疫组织化学方法检测PLD 1和PKCα在肢端雀斑样黑色素瘤（ALM）和浅表扩散性黑色素瘤（SSM）原发灶和转移灶中的表达。此外，使用腺病毒介导的基因转移技术在人黑素瘤细胞系HM 3 KO中检测PKCα对PLD 1的调节机制。PLD 1和DKCα在原发灶和转移灶中均呈强阳性表达。相反，在ALM病变中，这两种蛋白的表达随着肿瘤的进展而显著增加; PLD 1和PKCα的表达在原发性ALM病变的放射状生长期几乎为阴性，而在晚期ALM病变中以进展相关的方式同步增加，包括垂直生长期，而在晚期ALM病变中，PLD 1和PKC α的表达则随着肿瘤的进展而显著增加。 ...更多信息 生长期和转移病灶。免疫沉淀研究表明，PLD 1和PKCα在静息黑色素瘤细胞中生理相关。在腺病毒介导的同时过表达PLD 1和PKCα或PLD 1和PKCα的激酶阴性突变体后，使用HM 3 KO细胞进行的进一步免疫沉淀研究显示，在没有外部信号的情况下，PKCα和PKCα的激酶阴性突变体都与黑素瘤细胞中的PLD 1相关。在黑色素瘤细胞中，通过腺病毒载体过表达PKCα或PKCα的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。此外，增强的基础PLD活性增加了HM 3 KO细胞的体外侵袭潜力。这些结果表明，PLD 1和PKCα的上调在ALM从径向生长期向垂直生长期的进展中起作用。目前的结果还表明，PKCα与PLD 1相关，并以蛋白磷酸化非依赖性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。少

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