Joutel A, Andreux F, Gaulis S, Domenga V, Cecillon M, Battail N, Piga N, Chapon F, Godfrain C, Tournier-Lasserve E
INSERM U25, Faculté de Médecine Necker-Enfants Malades, Paris 75730, France. Laboratoire de Cytogénétique, Hôpital Lariboisière, Paris 75010, France.
J Clin Invest. 2000 Mar;105(5):597-605. doi: 10.1172/JCI8047.
Mutations in Notch3 cause CADASIL (cerebral autosomal dominant adult onset arteriopathy), which leads to stroke and dementia in humans. CADASIL arteriopathy is characterized by major alterations of vascular smooth muscle cells and the presence of specific granular osmiophilic deposits. Patients carry highly stereotyped mutations that lead to an odd number of cysteine residues within EGF-like repeats of the Notch3 receptor extracellular domain. Such mutations may alter the processing or the trafficking of this receptor, or may favor its oligomerization. In this study, we examined the Notch3 expression pattern in normal tissues and investigated the consequences of mutations on Notch3 expression in transfected cells and CADASIL brains. In normal tissues, Notch3 expression is restricted to vascular smooth muscle cells. Notch3 undergoes a proteolytic cleavage leading to a 210-kDa extracellular fragment and a 97-kDa intracellular fragment. In CADASIL brains, we found evidence of a dramatic and selective accumulation of the 210-kDa Notch3 cleavage product. Notch3 accumulates at the cytoplasmic membrane of vascular smooth muscle cells, in close vicinity to but not within the granular osmiophilic material. These results strongly suggest that CADASIL mutations specifically impair the clearance of the Notch3 ectodomain, but not the cytosolic domain, from the cell surface.
Notch3基因的突变会导致大脑常染色体显性遗传性成人发病动脉病（CADASIL），进而引发人类中风和痴呆。CADASIL动脉病的特征是血管平滑肌细胞发生重大改变，并存在特定的嗜锇颗粒沉积。患者携带高度定型的突变，这些突变导致Notch3受体细胞外结构域的表皮生长因子（EGF）样重复序列中半胱氨酸残基数量为奇数。此类突变可能会改变该受体的加工或运输过程，或者有利于其寡聚化。在本研究中，我们检测了正常组织中Notch3的表达模式，并研究了突变对转染细胞和CADASIL患者大脑中Notch3表达的影响。在正常组织中，Notch3的表达仅限于血管平滑肌细胞。Notch3会发生蛋白水解切割，产生一个210 kDa的细胞外片段和一个97 kDa的细胞内片段。在CADASIL患者的大脑中，我们发现有证据表明210 kDa的Notch3切割产物出现显著且选择性的积累。Notch3在血管平滑肌细胞的细胞质膜处积累，紧邻嗜锇颗粒物质但不在其内部。这些结果有力地表明，CADASIL突变特异性地损害了Notch3胞外结构域从细胞表面的清除，而不是胞质结构域的清除。
