The ectodomain of the Notch3 receptor accumulates within the cerebrovasculature of CADASIL patients

Mutations in Notch3 cause CADASIL (cerebral autosomal dominant adult onset arteriopathy), which leads to stroke and dementia in humans. CADASIL arteri opathy is characterized by major alterations of vascular smooth muscle cell s and the presence of specific granular osmiophilic deposits. Patients carr y highly stereotyped mutations that lead to an odd number of cysteine resid ues within EGF-like repeats of the Notch3 receptor extracellular domain. Su ch mutations may alter the processing or the trafficking of this receptor, or may favor its oligomerization. In this study, we examined the Notch3 exp ression pattern in normal tissues and investigated the consequences of muta tions on Notch3 expression in transfected cells and CADASIL brains. In norm al tissues, Notch3 expression is restricted to vascular smooth muscle cells . Notch3 undergoes a proteolytic cleavage leading to a 210-kDa extracellula r fragment and a 97-kDa intracellular fragment. In CADASIL brains, we found evidence of a dramatic and selective accumulation of the 210-kDa Notch3 cl eavage product. Notch3 accumulates at the cytoplasmic membrane of vascular smooth muscle cells, in close vicinity to but not within the granular osmio philic material. These results strongly suggest that CADASIL mutations spec ifically impair the clearance of the Notch3 ectodomain, but not the cytosol ic domain, from the cell surface.