Gain-of-function mutations in a member of the Src family kinases cause autoinflammatory bone disease in mice and humans
Gain-of-function mutations in a member of the Src family kinases cause autoinflammatory bone disease in mice and humans
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DOI：
10.1073/pnas.1819825116
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发表时间：
2019-06-11
影响因子：
11.1
通讯作者：
de Angelis, Martin Hrabe
中科院分区：
文献类型：
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作者：
Abe, Koichiro;Cox, Allison;de Angelis, Martin Hrabe
自身炎性综合征的特征在于先天免疫应答失调，随后发生急性自发性炎症。慢性复发性多灶性骨髓炎（CRMO）是一种自身炎症性骨病，表现为骨痛和局部肿胀。从诱变筛选中分离的Ali 18小鼠表现出自发性炎性爪表型，包括无菌骨髓炎和全身性骨矿物质密度降低。为了阐明该疾病的分子基础，试图定位克隆Ali 18的致病基因。利用候选基因的方法，在C-末端区域的Fgr，Src家族酪氨酸激酶（SFKs）的成员，错义突变，被确定。为了功能确认，通过CRISPR/Cas9介导的基因组编辑在Ali 18小鼠中引入Fgr的N末端处的额外突变。Fgr的N-末端有害突变消除了Ali 18小鼠的炎性表型，但同一区域的框内和错义突变继续表现出表型。Fgr无效突变小鼠在形态学上正常的事实表明，该模型中的炎症依赖于Fgr产物。此外，与野生型Fgr相比，Fgr（Ali 18）的C-末端负调节磷酸化水平明显降低。此外，对99名CRMO患者（包括88名三人组（先证者和父母））进行的全外显子组测序确定了13名FGR杂合编码序列变异的患者，包括两种影响激酶活性的错义突变蛋白。我们的研究结果有力地表明，Fgr的功能获得性突变参与无菌性骨髓炎，因此，使用特异性抑制剂靶向SFKs可以有效治疗该疾病。
Autoinflammatory syndromes are characterized by dysregulation of the innate immune response with subsequent episodes of acute spontaneous inflammation. Chronic recurrent multifocal osteomyelitis (CRMO) is an autoinflammatory bone disorder that presents with bone pain and localized swelling. Ali18 mice, isolated from a mutagenesis screen, exhibit a spontaneous inflammatory paw phenotype that includes sterile osteomyelitis and systemic reduced bone mineral density. To elucidate the molecular basis of the disease, positional cloning of the causative gene for Ali18 was attempted. Using a candidate gene approach, a missense mutation in the C-terminal region of Fgr, a member of Src family tyrosine kinases (SFKs), was identified. For functional confirmation, additional mutations at the N terminus of Fgr were introduced in Ali18 mice by CRISPR/Cas9-mediated genome editing. N-terminal deleterious mutations of Fgr abolished the inflammatory phenotype in Ali18 mice, but in-frame and missense mutations in the same region continue to exhibit the phenotype. The fact that Fgr null mutant mice are morphologically normal suggests that the inflammation in this model depends on Fgr products. Furthermore, the levels of C-terminal negative regulatory phosphorylation of Fgr(Ali18) are distinctly reduced compared with that of wild-type Fgr. In addition, whole-exome sequencing of 99 CRMO patients including 88 trios (proband and parents) identified 13 patients with heterozygous coding sequence variants in FGR, including two missense mutant proteins that affect kinase activity. Our results strongly indicate that gain-of-function mutations in Fgr are involved in sterile osteomyelitis, and thus targeting SFKs using specific inhibitors may allow for efficient treatment of the disease.