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

Kit和Kit配体(RI)编码基因在黑素母细胞分化过程中起着重要作用。我们以前建立了三个永生但不同的小鼠神经脊(NC)细胞群体。NCCmelb4M5细胞不表达Kit并独立于KL生长；它们有分化为NCCmelb4细胞的潜力，NCCmelb4细胞是Kit阳性的黑素细胞前体。NCCmer5细胞具有分化黑素细胞的特征。三种细胞系均表达BMP受体。BMP-4上调大多数未成熟NCCmelb4M5细胞Kit蛋白和mRNA的表达。BMP-4拮抗剂noggin可显著降低BMP-4诱导的Kit表达。Western印迹分析显示外源性BMP-4可导致NCCmelb4M5细胞中Smads的磷酸化。结果表明，BMP-4能激活Kit启动子在NCCmelb4M5细胞中的表达。我们得出结论，BMP-4是活性的，并参与了大多数未成熟黑素细胞前体细胞Kit表达的调节。我们进一步研究了BMP-4对体外培养的野生型小鼠NC细胞的影响。与稀释液处理的对照组相比，在培养液中添加BMP-4可以增加Kit阳性细胞的数量。我们已经确定BMP-4是产前Kit阴性的黑素母细胞在进入Kit依赖的黑素生成周期之前的一个重要因素。

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.