A CADASIL-mutated Notch 3 receptor exhibits impaired intracellular trafficking and maturation but normal ligand-induced signaling
A CADASIL-mutated Notch 3 receptor exhibits impaired intracellular trafficking and maturation but normal ligand-induced signaling
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DOI：
10.1073/pnas.252624099
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发表时间：
2002-12-24
影响因子：
11.1
通讯作者：
Lundkvist, J
中科院分区：
文献类型：
--
作者：
Karlström, H;Beatus, P;Lundkvist, J
Notch受体是单跨膜受体，其胞外域含有大量表皮生长因子样重复序列（EGF重复序列）。人类 Notch 3 受体 EGF 重复序列的突变会导致血管性痴呆疾病：伴有皮质下梗死和白质脑病的常染色体显性遗传性脑动脉病 (CADASIL)。绝大多数 CADASIL 突变是错义突变，在 EGF 重复序列中去除或插入半胱氨酸残基，但尚不清楚这些突变是否主要影响受体运输、成熟和/或信号传导。为了解决这个问题，我们生成并分析了表达野生型小鼠 Notch 3 (mNotch 3) 或突变型 mNotch 3(R142C) 的稳定细胞系，突变型 mNotch 3(R142C) 对应于人类中常见的 CADASIL 形式的 Notch 3、Notch 3(R141C)。我们发现，与野生型 mNotch 3 相比，mNotch 3(R142C) 在位点 1 切割构型中表达的比例较低，并且细胞表面出现的 mNotch 3(R142C) 量减少。这一观察结果伴随着 mNotch 3(R142Cd) 形成细胞内聚集体的较高倾向，这可能是由于 mNotch 3(R142C) 形成细胞内聚集体的倾向增加。 分泌途径中的积累或运输减慢。与受损的细胞表面表达相反，mNotch 3(R142C) 对 Delta 1 和 Jagged 1 的信号反应与野生型 mNotch 3 一样好。总而言之，这些数据表明 mNotch 3 的运输和定位而不是信号传导在 mNotch 3(R142C) 中受到影响。
Notch receptors are single transmembrane receptors that contain a large number of epidermal growth factor-like repeats (EGF repeats) in their extracellular domains. Mutations in the EGF repeats of the human Notch 3 receptor lead to the vascular dementia disease Cerebral Autosomal Dominant Arteriopathy with Subcortical Infarcts and Leukoencephalopathy (CADASIL). The vast majority of CADASIL mutations are missense mutations removing or inserting cysteine residues in the EGF repeats, but it is not yet clear whether these mutations primarily affect receptor trafficking, maturation, and/or signaling. To address this issue, we have generated and analyzed stable cell lines expressing either wildtype murine Notch 3 (mNotch 3) or the mutant mNotch 3(R142C), which corresponds to the prevalent CADASIL form of Notch 3, Notch 3(R141C) in humans. We find that a lower proportion of mNotch 3(R142C) is expressed in the site 1-cleaved configuration, and that reduced amounts of mNotch 3(R142C) appear at the cell surface, as compared with wild-type mNotch 3. This observation is accompanied by a higher propensity for mNotch 3(R142Cd) to form intracellular aggregates, which may be a result of increased accumulation or slowed transport in the secretory pathway. In contrast to the impaired cell surface expression, mNotch 3(R142C) signals equally well in response to Delta 1 and Jagged 1 as wild-type mNotch 3. Taken together, these data suggest that trafficking and localization rather than signaling of mNotch 3 are affected in mNotch 3(R142C).