A novel cysteine-sparing G73A mutation of NOTCH3 in a Chinese CADASIL family

  • Liyan Huang
  • Wei Li
  • Yi Li
  • Chaoyuan Song
  • Pin Wang
  • Hongchun Wang
  • Xiulian SunEmail author
Original Article


Cerebral autosomal dominant arteriopathy with subcortical infarcts and leukoencephalopathy (CADASIL) is the most common monogenic disease leading to stroke and vascular dementia. CADASIL is an inherited small blood vessel disease caused by mutations in the gene encoding the neurogenic locus notch homolog protein 3 (NOTCH3). NOTCH3 is large type I membrane receptor mainly expressed in vascular smooth muscle cells and pericytes. Most identified mutations result in insert or deletion of a cysteine residue within the EGF-like repeats. To date, some cases with a cysteine-sparing mutant have been described. Genetic analysis revealed a novel mutation in NOTCH3 in a CADASIL family. Molecular analysis revealed its potential pathogenic mechanism in causing CADASIL. In this paper, we present a Chinese family with a novel cysteine-sparing mutation in exon 3 (c.218G>C, p.G73A) of the NOTCH3 gene. Family carriers of the same mutation presented with symptoms and imaging abnormalities characteristic of CADASIL. The location of glycine 73 in between C5-C6 disulfide bond of EGF-like domain 1 shows high conservation from humans to zebra fish. It has previously been suggested that the aggregate-prone property of mutant NOTCH3 contributes to a cytotoxic effect in the pathogenic mechanism underlying CADASIL. Here, we investigated the pathogenic mechanism of the new mutation in vitro using HEK293 cells transfected with either a wild-type (WT) or c.218G>C (p.G73A) NOTCH3ECD plasmids, and we found p.G73A NOTCH3ECD was more prone to form aggregation and resistant to degradation. Moreover, the p.G73A NOTCH3ECD compromised cell viability by promoting apoptosis. Two known CADASIL mutants R133C and R75P showed similar results with G73A mutants. Our study here identified G73A as a new mutation in NOTCH3 to cause CADASIL and revealed that the G73A mutation and two known mutants R75P and R133C decreased NOTCH3 protein turnover and induced cell death.


CADASIL NOTCH3 Apoptosis Stroke Dementia 



analysis of variance


cerebral autosomal dominant arteriopathy with subcortical infarcts and leukoencephalopathy


Cell Counting Kit-8


coronary heart disease






diabetes mellitus




extracellular domain


extracellular matrix

EGF-like repeats

epidermal growth factor-like repeats


endoplasmic reticulum


flow cytometry


fluorescein isothiocyanate


fluid-attenuated inversion recovery magnetic resonance imaging


granular osmiophilic material


Hamilton Anxiety Scale


human embryonic kidney 293 cells


intracellular domain


impaired glucose tolerance


magnetic resonance imaging


mini-mental state examination


neurogenic locus notch homolog protein 3


phosphate-buffered saline


propidium iodide


susceptibility weighted imaging


unfolded protein response


vascular smooth muscle cells


T2-weighted white matter hyperintensity


wild type



We thank Dr. Weihong Song for his instructive suggestions.

Availability of data and materials

All data generated or analyzed during this study are included in this published article (and as supplement original data.pdf).

Author’s contributions

L. H. and X. S. designed the experiments and wrote the manuscript. W. L. collected the clinical data. L. H. performed the molecular studies. Y. L., C. S., P. W., and H. W. helped in preparing the materials. All authors read and approved the final manuscript.

Funding information

This study was supported by grants from NSFC (81771155) to Xiulian Sun and grants from Shandong Key Research Project (2018GSF118104) to Hongchun Wang.

Compliance with ethical standards

Ethical statements

Study protocols were approved by the Medical Ethical Committee of Qingdao Municipal Hospital (2016-002). All persons gave their informed consent prior to their inclusion in the study.

Consent for publication

Not applicable.

Competing interests

The authors declare that they have no conflict of interest.


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Copyright information

© Springer-Verlag GmbH Germany, part of Springer Nature 2019

Authors and Affiliations

  • Liyan Huang
    • 1
  • Wei Li
    • 1
    • 3
  • Yi Li
    • 1
  • Chaoyuan Song
    • 5
  • Pin Wang
    • 4
  • Hongchun Wang
    • 6
  • Xiulian Sun
    • 2
    Email author
  1. 1.Department of NeurologyQilu Hospital of Shandong UniversityJinanChina
  2. 2.Brain Research InstituteQilu Hospital of Shandong UniversityJinanChina
  3. 3.Department of NeurologyQingdao Municipal HospitalQingdaoChina
  4. 4.Otolarygology Key Lab of National Health CommitteeQilu Hospital of Shandong UniversityJinanChina
  5. 5.Department of NeurologySecondary Hospital of Shandong UniversityJinanChina
  6. 6.Department of Clinical LaboratoryQilu Hospital of Shandong UniversityJinanChina

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