Xu Daosong, Sharma Chandan, Hemler Martin E
Dana-Farber Cancer Institute and Department of Pathology, Harvard Medical School, Boston, Massachusetts 02115, USA.
FASEB J. 2009 Nov;23(11):3674-81. doi: 10.1096/fj.09-133462. Epub 2009 Jul 8.
Using mass spectrometry, we identified ADAM10 (a membrane-associated metalloproteinase) as a partner for TSPAN12, a tetraspanin protein. TSPAN12-ADAM10 interaction was confirmed by reciprocal coimmunoprecipitation in multiple tumor cell lines. TSPAN12, to a greater extent than other tetraspanins (CD81, CD151, CD9, and CD82), associated with ADAM10 but not with ADAM17. Overexpression of TSPAN12 enhanced ADAM10-dependent shedding of amyloid precursor protein (APP) in MCF7 (breast cancer) and SH-SY5Y (neuroblastoma) cell lines. Conversely, siRNA ablation of endogenous TSPAN12 markedly diminished APP proteolysis in both cell lines. Furthermore, TSPAN12 overexpression enhanced ADAM10 prodomain maturation, whereas TSPAN12 ablation diminished ADAM10 maturation. A palmitoylation-deficient TSPAN12 mutant failed to associate with ADAM10, inhibited ADAM10-dependent proteolysis of APP, and inhibited ADAM10 maturation, most likely by interfering with endogenous wild-type TSPAN12. In conclusion, TSPAN12 serves as a novel and robust partner for ADAM10 and promotes ADAM10 maturation, thereby facilitating ADAM10-dependent proteolysis of APP. This novel mode of regulating APP cleavage is of relevance to Alzheimer's disease therapy.
利用质谱分析法，我们鉴定出ADAM10（一种膜相关金属蛋白酶）是四跨膜蛋白TSPAN12的相互作用蛋白。通过在多种肿瘤细胞系中进行相互免疫共沉淀，证实了TSPAN12与ADAM10之间的相互作用。与其他四跨膜蛋白（CD81、CD151、CD9和CD82）相比，TSPAN12与ADAM10的结合更为紧密，但不与ADAM17结合。在MCF7（乳腺癌）和SH-SY5Y（神经母细胞瘤）细胞系中，TSPAN12的过表达增强了ADAM10依赖性的淀粉样前体蛋白（APP）的裂解。相反，内源性TSPAN12的siRNA敲除显著减少了这两种细胞系中APP的蛋白水解。此外，TSPAN12的过表达增强了ADAM10前结构域的成熟，而TSPAN12的敲除则减少了ADAM10的成熟。一种棕榈酰化缺陷的TSPAN12突变体无法与ADAM10结合，抑制了ADAM10依赖性的APP蛋白水解，并抑制了ADAM10的成熟，这很可能是通过干扰内源性野生型TSPAN12实现的。总之，TSPAN12是ADAM10一种新的、强有力的相互作用蛋白，并促进ADAM10的成熟，从而促进ADAM10依赖性的APP蛋白水解。这种调节APP裂解的新模式与阿尔茨海默病的治疗相关。
