Laboratory of Osteoblast Biology and Pathology, Inserm U606, Paris, France.
Hum Mol Genet. 2010 May 1;19(9):1678-89. doi: 10.1093/hmg/ddq045. Epub 2010 Feb 2.
Dysregulations of osteoblast function induced by gain-of-function genetic mutations in fibroblast growth factor receptors (FGFRs) cause premature fusion of cranial sutures in syndromic craniosynostosis. The pathogenic signaling mechanisms induced by FGFR genetic mutations in human craniosynostosis remain largely unknown. In this study, we have used microarray analysis to investigate the signaling pathways that are activated by FGFR2 mutations in Apert craniosynostosis. Transcriptomic analysis revealed that EGFR and PDGFRalpha expression is abnormally increased in human Apert calvaria osteoblasts compared with wild-type cells. Quantitative RT-PCR and western blot analyses in Apert osteoblasts and immunohistochemical analysis of Apert sutures confirmed the increased EGFR and PDGFRalpha expression in vitro and in vivo. We demonstrate that pharmacological inhibition of EGFR and PDGFR reduces the pathological upregulation of phenotypic osteoblast genes and in vitro matrix mineralization in Apert osteoblasts. Investigation of the underlying molecular mechanisms revealed that activated FGFR2 enhances EGFR and PDGFRalpha mRNA expression via activation of PKCalpha-dependent AP-1 transcriptional activity. We also show that the increased EGFR protein expression in Apert osteoblasts results in part from a post-transcriptional mechanism involving increased Sprouty2-Cbl interaction, leading to Cbl sequestration and reduced EGFR ubiquitination. These data reveal novel molecular crosstalks between activated FGFR2, EGFR and PDGFRalpha that functionally contribute to the osteoblastic dysfunction in Apert craniosynostosis, which may provide a molecular basis for novel therapeutic approaches in this severe skeletal disorder.
成纤维细胞生长因子受体（FGFRs）功能获得性基因突变引起的成骨细胞功能失调导致综合征性颅缝早闭的颅缝过早融合。FGFR 基因突变在人类颅缝早闭中诱导的致病信号机制在很大程度上尚不清楚。在这项研究中，我们使用微阵列分析研究了 FGFR2 突变在 Apert 颅缝早闭中激活的信号通路。转录组分析显示，与野生型细胞相比，Apert 颅骨成骨细胞中 EGFR 和 PDGFRalpha 的表达异常增加。Apert 成骨细胞中的定量 RT-PCR 和 Western blot 分析以及 Apert 缝的免疫组织化学分析证实了体外和体内 EGFR 和 PDGFRalpha 表达的增加。我们证明，EGFR 和 PDGFR 的药理学抑制可降低 Apert 成骨细胞中病理性上调的表型成骨细胞基因和体外基质矿化。对潜在分子机制的研究表明，激活的 FGFR2 通过激活依赖 PKCalpha 的 AP-1 转录活性增强 EGFR 和 PDGFRalpha mRNA 表达。我们还表明，Apert 成骨细胞中 EGFR 蛋白表达的增加部分是由于涉及增加 Sprouty2-Cbl 相互作用的转录后机制，导致 Cbl 隔离和减少 EGFR 泛素化。这些数据揭示了激活的 FGFR2、EGFR 和 PDGFRalpha 之间的新的分子串扰，这些串扰在功能上有助于 Apert 颅缝早闭中成骨细胞功能障碍，这可能为这种严重骨骼疾病的新治疗方法提供分子基础。
