Novel variants of muscle calpain 3 identified in human melanoma cells: cisplatin-induced changes in vitro and differential expression in melanocytic lesions
Novel variants of muscle calpain 3 identified in human melanoma cells: cisplatin-induced changes in vitro and differential expression in melanocytic lesions
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DOI：
10.1093/carcin/bgp098
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发表时间：
2009-06-01
期刊：
影响因子：
4.7
通讯作者：
Maellaro, E.
中科院分区：
文献类型：
--
作者：
Moretti, D.;Del Bello, B.;Maellaro, E.
钙蛋白酶是半胱氨酸蛋白酶，包括在人体组织中普遍表达的成员以及其他组织特异性异构体。钙蛋白酶3（p94）是一种肌肉特异性异构体，包含三个特殊序列（NS、IS1和IS2），其改变与2A型肢带型肌营养不良症密切相关，在该疾病中已记录到肌核凋亡。我们最近证明了普遍存在的钙蛋白酶在药物诱导的黑色素瘤细胞凋亡中具有促凋亡作用，这促使我们研究钙蛋白酶3在经历凋亡的人黑色素瘤细胞系以及黑素细胞病变中的表达。在黑色素瘤细胞系中，我们鉴定出了钙蛋白酶3的两种新型剪接变体（hMp78和hMp84）：它们具有一个非典型起始外显子和一个假定的核定位信号，较短的变体缺少IS1插入序列，并且这两种蛋白质都极不稳定。实际上，这两种异构体（主要以裂解形式存在）都定位于细胞质和核仁中。在顺铂处理的预凋亡细胞中，观察到新型变体的转录和自蛋白水解裂解都增加；普遍存在的钙蛋白酶抑制剂钙肽素可阻止后一事件，它也能够防止细胞凋亡。有趣的是，在黑素细胞病变中，与良性痣相比，这些新型变体在最具侵袭性的病变中，即垂直生长期黑色素瘤以及更具侵袭性且通常对凋亡高度抗性的转移性黑色素瘤细胞中的表达显著下调。总体而言，我们的观察结果表明，钙蛋白酶3变体在黑色素瘤细胞中可发挥促凋亡作用，并且如在高度侵袭性病变中所观察到的那样，其下调可能有助于黑色素瘤的进展。
Calpains are cysteine proteases comprising members ubiquitously expressed in human tissues and other tissue-specific isoforms. Alterations of calpain 3 (p94), the muscle-specific isoform that contains three peculiar sequences (NS, IS1 and IS2), are strictly associated to the limb-girdle muscular dystrophy type 2A, in which a myonuclear apoptosis has been documented. Our recent demonstration of a proapoptotic role of ubiquitous calpains in drug-induced apoptosis of melanoma cells prompted us to investigate the expression of calpain 3 in human melanoma cell lines undergoing apoptosis and in melanocytic lesions. In melanoma cell lines, we have identified two novel splicing variants of calpain 3 (hMp78 and hMp84): they have an atypical initiation exon and a putative nuclear localization signal, the shorter one lacks IS1 inset and both proteins are extremely unstable. Virtually, both isoforms (prevalently as cleavage forms) are localized in cytoplasm and in nucleoli. In cisplatin-treated preapoptotic cells, an increase of both transcription and autoproteolytic cleavage of the novel variants is observed; the latter event is prevented by the inhibitor of ubiquitous calpains, calpeptin, which is also able to protect from apoptosis. Interestingly, among melanocytic lesions, the expression of these novel variants is significantly downregulated, compared with benign nevi, in the most aggressive ones, i.e. in vertical growth phase melanoma and, even more, in metastatic melanoma cells, characterized by invasiveness properties and usually highly resistant to apoptosis. On the whole, our observations suggest that calpain 3 variants can play a proapoptotic role in melanoma cells and its downregulation, as observed in highly aggressive lesions, could contribute to melanoma progression.