FAM20A mutations and transcriptome analyses of dental pulp tissues of enamel renal syndrome.
FAM20A mutations and transcriptome analyses of dental pulp tissues of enamel renal syndrome.
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釉质肾综合征牙髓组织 FAM20A 突变及转录组分析
DOI：
10.1111/iej.13928
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发表时间：
2023
影响因子：
5
通讯作者：
Hu,JanC-C
中科院分区：
文献类型：
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作者：
Wang,Shih-Kai;Zhang,Hong;Wang,Yin-Lin;Lin,Hung-Ying;Seymen,Figen;Koruyucu,Mine;Wright,JTimothy;Kim,Jung-Wook;Simmer,JamesP;Hu,JanC-C
目的双等位基因功能丧失 FAM20 突变导致 IG 型釉质生成不全 (AI)，即牙釉质肾综合征 (ERS)，其特征为严重牙釉质发育不全、牙齿萌出延迟/失败、牙髓内钙化、牙龈增生和肾钙质沉着症。 FAM20A 与 FAM20C（高尔基体酪蛋白激酶 (GCK)）结合，增强其磷酸化对生物矿化至关重要的分泌蛋白的功能。虽然许多FAM20非致病性突变已被报道，但ERS中口腔牙齿异常的发病机制仍有待阐明。本研究旨在鉴定 ERS 表型患者的致病突变，并揭示 ERS 牙髓内钙化的分子机制。方法学对 8 个家庭和 2 例 AI 发育不全的散发病例进行表型表征和全外显子组分析。进行小基因测定以研究 aFAM20Aplice 位点变异的分子后果。对 ERS 和对照的牙髓组织进行 RNA 测序，然后进行转录谱分析和基因本体 (GO) 分析。结果每个受影响个体都证实了 BiallelicFAM20 突变，包括 7 个新的致病变异：c.590‐5T>A、c.625T>A (p.Cys209Ser)、c.771del (p.Gln258Argfs*28)、 c.832_835delinsTGTCCGACGGTGTCCGACGGTGTC CA (p.Val278Cysfs*29)、c.1232G>A (p.Arg411Gln)、c.1297A>G (p.Arg433Gly) 和 c.1351del (p.Gln451Serfs*4)。 c.590-5T>A 剪接位点突变导致外显子 3 跳跃，从而导致 FAM20A 蛋白 p.(Asp197_Ile214delinsVal) 的独特区域的框内删除。对ERS牙髓组织中差异表达基因的分析表明，参与生物矿化，特别是牙本质形成的基因显着上调，如DSPP、MMP9、MMP20和WNT10A。富集分析表明与 BMP 和 SMAD 信号通路相关的基因集过多。相比之下，与炎症和轴突发育相关的 GO 术语代表性不足。 BMP信号基因中，BMP激动剂GDF7、GDF15、BMP3、BMP8A、BMP8B、BMP4和BMP6在ERS牙髓组织中表达上调，而BMP拮抗剂GREM1、BMPER和VWC2在ERS牙髓组织中表达降低。结论BMP信号上调是ERS牙髓内钙化的基础。 FAM20A 在牙髓组织稳态和预防软组织异位矿化中发挥重要作用。这一关键功能可能取决于 MGP（基质 Gla 蛋白），这是一种有效的矿化抑制剂，必须被 FAM20A-FAM20C 激酶复合物正确磷酸化。
AimBiallelic loss‐of‐functionFAM20Amutations cause amelogenesis imperfecta (AI) type IG, better known as enamel renal syndrome (ERS), characterized by severe enamel hypoplasia, delayed/failed tooth eruption, intrapulpal calcifications, gingival hyperplasia and nephrocalcinosis. FAM20A binds to FAM20C, the Golgi casein kinase (GCK) and potentiates its function to phosphorylate secreted proteins critical for biomineralization. While manyFAM20Apathogenic mutations have been reported, the pathogeneses of orodental anomalies in ERS remain to be elucidated. This study aimed to identify disease‐causing mutations for patients with ERS phenotypes and to discern the molecular mechanism underlying ERS intrapulpal calcifications.MethodologyPhenotypic characterization and whole exome analyses were conducted for 8 families and 2 sporadic cases with hypoplastic AI. A minigene assay was performed to investigate the molecular consequences of aFAM20Asplice‐site variant. RNA sequencing followed by transcription profiling and gene ontology (GO) analyses were carried out for dental pulp tissues of ERS and the control.ResultsBiallelicFAM20Amutations were demonstrated for each affected individual, including 7 novel pathogenic variants: c.590‐5T>A, c.625T>A (p.Cys209Ser), c.771del (p.Gln258Argfs*28), c.832_835delinsTGTCCGACGGTGTCCGACGGTGTC CA (p.Val278Cysfs*29), c.1232G>A (p.Arg411Gln), c.1297A>G (p.Arg433Gly) and c.1351del (p.Gln451Serfs*4). The c.590‐5T>A splice‐site mutation caused Exon 3 skipping, which resulted in an in‐frame deletion of a unique region of the FAM20A protein, p.(Asp197_Ile214delinsVal). Analyses of differentially expressed genes in ERS pulp tissues demonstrated that genes involved in biomineralization, particularly dentinogenesis, were significantly upregulated, such asDSPP,MMP9,MMP20andWNT10A. Enrichment analyses indicated overrepresentation of gene sets associated with BMP and SMAD signalling pathways. In contrast, GO terms related to inflammation and axon development were underrepresented. Among BMP signalling genes, BMP agonistsGDF7,GDF15,BMP3,BMP8A,BMP8B,BMP4andBMP6were upregulated, while BMP antagonistsGREM1,BMPERandVWC2showed decreased expression in ERS dental pulp tissues.ConclusionsUpregulation of BMP signalling underlies intrapulpal calcifications in ERS. FAM20A plays an essential role in pulp tissue homeostasis and prevention of ectopic mineralization in soft tissues. This critical function probably depends upon MGP (matrix Gla protein), a potent mineralization inhibitor that must be properly phosphorylated by FAM20A‐FAM20C kinase complex.