The role of BMP and Kit in melanocyte and melanoma cell differentiation

编码Kit和Kit配体（RI）的基因在成黑素细胞的分化中起重要作用。我们以前建立了三个不朽的，但不同的细胞群的小鼠神经嵴（NC）细胞。NCCmelb 4 M5细胞不表达Kit并且独立于KL生长;它们具有分化成NCCmelb 4细胞的潜力，NCCmelb 4细胞是Kit阳性黑素细胞前体。NCCmelan 5细胞显示出分化的黑素细胞的特征。所有三种细胞系均表现出BMP受体表达。BMP-4在大多数未成熟的NCCmelb 4 M5细胞中上调Kit蛋白和mRNA的表达。BMP-4拮抗剂Noggin显著降低BMP-4诱导的Kit表达。Western印迹分析显示，外源性BMP-4导致NCCMelb 4 M5细胞中Smads的磷酸化。利用转染的Kit启动子报告基因，我们发现BMP-4可以激活转染的NCCmelb 4 M5细胞中的Kit启动子。我们的结论是，BMP-4是活跃的，并参与大多数未成熟的黑素细胞前体的Kit表达的调节。我们使用从野生型小鼠培养的原代NC细胞进一步研究了BMP-4在体外的影响。与稀释液处理的对照相比，向培养基中添加BMP-4增加了Kit阳性细胞的数量。我们已经确定BMP-4作为一个重要的因素，产前试剂盒阴性黑素细胞之前，进入试剂盒依赖性周期的黑素生成。

Genes encoding Kit and the Kit ligand (RI) play essential roles in the differentiation of melanoblasts. We previously established three immortal but distinct cell populations of mouse neural crest (NC) cells. NCCmelb4M5 cells do not express Kit and grow independently of KL; they have the potential to differentiate into NCCmelb4 cells, which are Kit-positive melanocyte precursors. NCCmelan5 cells show the characteristics of differentiated melanocytes. All three cell lines demonstrated BMP-receptor expression. BMP-4 upregulated Kit protein and mRNA expression in most immature NCCmelb4M5 cells. Noggin, a BMP-4 antagonist, dramatically decreased the Kit expression induced by BMP-4. Western blot analysis revealed that extrinsic BMP-4 leads to the phosphorylation of Smads in NCCmelb4M5 cells. Using transfected Kit-promoter reporter we showed BMP-4 could activate Kit promoter in transfected NCCmelb4M5 cells. We conclude that BMP-4 is active and is involved in the regulation of Kit expression on most immature melanocyte precursors. We further investigated the influence of BMP-4 in vitro using primary NC cells cultured from wild-type mice. Addition of BMP-4 to the medium increased the number of Kit-positive cells compared to diluent-treated controls. We have identified BMP-4 as an important factor for prenatal Kit-negative melanoblasts just prior to entering the Kit-dependent cycle of melanogenesis.