家族性脑海绵状血管畸形（FCCM）是一种具有不完全外显率的常染色体显性遗传病，可导致出血、癫痫或神经功能缺损。目前FCCM的致病基因定位于CCM1/KRIT1、CCM2/MGC4607、CCM3/PDCD10三个片段，但突变位点具有显著异质性且汉族人群研究较少。前期工作中，我们收集了大宗汉族FCCM家系，报道了多个新的无义、缺失、剪接受体位点突变，均位于CCM1/KRIT1基因上，但这些突变对KRIT1蛋白功能的影响尚不明确。而且，生物信息学分析提示这些突变除影响编码潜能之外，还可能影响mRNA的剪接，包括剪接位点识别异常和剪接调控异常，从而导致外显子跳跃。本研究首先利用minigene剪接报告系统和RNA pull-down技术探讨这些突变导致mRNA异常剪接的机制，进而构建体外真核表达体系，在细胞水平验证这些突变对KRIT1蛋白功能的影响，将为FCCM的致病机制和基因诊断提供重要依据。

Familial cerebral cavernous malformation (FCCM) is an autosomal-dominant disease with incomplete penetrance, clinically manifesting as hemorrhage, epilepsy and neurological deficits. The pathogenic genes of FCCM have been located in three chromosomal fragments: CCM1/KRIT1, CCM2/MGC4607 and CCM3/PDCD10; nevertheless, the mutations exhibit considerable heterogeneity and there is a paucity of relevant studies in the Chinese population. In our previous work, we collected a large-sample Chinese FCCM families and reported several new mutations, including nonsense, deletion and splicing-acceptor-site variants. All these mutations are located in the CCM1/KRIT1 gene, while the effects of these mutations on the function of KRIT1 protein remain unclear. Moreover, bioinformatic analyses indicate that these mutations can not only affect the coding potential, but leading to aberrant splicing as well. The underlying mechanisms include abnormal splicing site recognition and abnormal splicing regulation, which may cause exon skipping. In this study, we intend to investigate the mechanism of mutation-related aberrant splicing by using minigene splicing reporting system and RNA pull-down methods. Then, we will construct an in-vitro eukaryotic expression system, in which we can validate the effects of mutations on the function of KRIT1 protein. This study is expected to provide crucial evidence for the pathogenic mechanism and genetic diagnosis of FCCM.