Mutations of RNF213 are responsible for sporadic cerebral cavernous malformation and lead to a mulberry-like cluster in zebrafish

Abstract

Although familial forms of cerebral cavernous malformation are mainly attributed to three CCM genes ( KRIT1, CCM2 and PDCD10), no mutation is identified in sporadic cerebral cavernous malformation cases with a unique lesion, indicating additional genes for sporadic cerebral cavernous malformation. To screen the candidate genes, we conducted whole exome sequencing in 31 sporadic cerebral cavernous malformation patients and 32 healthy controls, and identified 5 affected individuals carrying 6 heterozygous deleterious mutations in RNF213 but no RNF213 mutation in healthy individuals. To further confirm RNF213 was associated with cerebral cavernous malformation, we generated rnf213a homozygous knockout zebrafish and found mutation of rnf213a in zebrafish led to a mulberry-like cluster of disordered-flow vascular channels which was reminiscent of human cerebral cavernous malformation. In addition, we revealed kbtbd7 and anxa6 were significantly downregulated due to rnf213a mutation through transcriptomic sequencing and RT-qPCR analysis. Based on the mulberry-like phenotype partly rescued by mRNA of kbtbd7 as well as anxa6, we suggested that rnf213a promoted mulberry-like cluster via downregulation of kbtbd7 and anxa6. Altogether, we firstly demonstrate RNF213is a novel candidate gene for sporadic cerebral cavernous malformation and the mutation of rnf213a is responsible for the mulberry-like cluster in zebrafish.

EZH2 Regulates ANXA6 Expression via H3K27me3 and Is Involved in Angiotensin II-Induced Vascular Smooth Muscle Cell Senescence

“…Proteome analysis of infrarenal abdominal aortic tissue and adjacent normal tissue from patients undergoing elective open AAA repair revealed 24 differentially expressed proteins, 10 of which were upregulated and 14 of which were downregulated (Figures 1(a) and 1(b)). The literature has showed that the differential expression of ANXA6 is associated with vascular disease [9][10][11]. Therefore, we chose the highly expressed ANXA6 for our study.…”

“…Between the membrane microdomains and the cytoskeleton, ANXA6 acts as a scaffold to function [ 8 ]. ANXA6 has been shown to be involved in sporadic cerebral cavernous malformations and mulberry poplar clusters in zebrafish [ 9 ]. At the same time, ANXA6 also promotes VSMC mineralization, thereby promoting vascular calcification [ 10 ].…”

A new syndrome of moyamoya disease, kidney dysplasia, aminotransferase elevation, and skin disease associated with de novo variants in RNF213

“…Though the exact function of RNF213 and the mechanism by which gene variants cause disease is unknown, in vitro and in vivo models support a role for RNF213 in angiogenesis. Specifically, abnormal RNF213 function in vitro causes abnormal cellular proliferation and dysregulation of inflammatory and extracellular matrix gene expression (Ohkubo et al, 2015 ), morpholino‐mediated and TALEN‐mediated suppression of rnf213 expression in zebrafish causes abnormal vascular sprouting (Lin et al, 2020 ; Liu et al, 2011 ; Wen et al, 2016 ), and murine models of rnf213 deficiency demonstrate abnormal angiogenesis and response to vascular injury (Ito et al, 2015 ; Sonobe et al, 2014 ). The exact mechanism by which variants in RNF213 disrupt angiogenesis and vascular injury response is unknown, but RNF213 is known to play a role in noncanonical Wnt and calcium signaling and NF‐kB pathway activation, which are pathways known to regulate growth and inflammatory signaling (Amal et al, 2019 ; Scholz et al, 2016 ; Takeda et al, 2020 ).…”

“…RNF213 encodes mysterin, a protein of unknown function, but with suspected E3 ubiquitin ligase and motor activity given its possession of a RING domain and two AAA+ ATPase domains (Morito et al, 2014 ). Though nearly ubiquitously expressed, RNF213 deficiency studies in murine and zebrafish models suggest a prominent role in angiogenesis and blood vessel response to injury and hypoxia, with the extra‐vascular function of RNF213 incompletely characterized (Ito et al, 2015 ; Lin et al, 2020 ; Liu et al, 2011 ; Sonobe et al, 2014 ; Wen et al, 2016 ).…”

Detailed phenotype ofRNF213p.R4810K variant identified by the Chinese patients with acute ischaemic stroke or transient ischaemic attack

“…The pathological mechanism underlying the effect of the RNF213 variant on abnormal angiogenesis remains unknown. Preclinical research has found that the loss of RNF213 function did not trigger the spontaneous development of smog-like vessels but caused irregular vascular wall formation as a critical regulator of cerebral endothelium integrity 22 36. According to research conducted on mice deficient in RNF213 , several metabolites have been found to be associated with the occurrence of abnormal angiogenesis, such as matrix metalloproteinase—9 and caveolin-1 37 38.…”