We herein report that deletion of mTOR in dental epithelia caused defective development of multiple cell layers of the enamel organ, which culminated in tooth malformation and cystogenesis. Specifically, cells of the stellate reticulum and stratum intermedium were poorly formed, resulting in cystic changes. The pre-ameloblasts failed to elongate along the apical-basal axis and persisted vigorous expression of Sox2 and P63, which are normally downregulated during cytodifferentiation. Expression of amelogenic markers was also attenuated in mutants. Cell proliferation and cell sizes in mutants were significantly reduced over time. Importantly, we found reduced amounts and aberrant aggregations of cytoskeletal components in mutants, along with attenuated expression of cytoskeleton regulator Cdc42, whose epithelial deletion causes a similar phenotype. Moreover, disruption of actin assembly in an organ culture system affected cell proliferation and cytodifferentiation of tooth germs, supporting a causative role of mTOR-regulated cytoskeleton dynamics for the observed phenotype of mTOR mutant mice. In further support of this view, we showed that mTOR overactivation caused increased cytoskeletal component synthesis and assembly, along with accelerated cytodifferentiation in the enamel organ. Finally, we demonstrated that mTOR regulated enamel organ development principally through the mTORC1 pathway.

我们在此报告，牙齿上皮细胞中mTOR的缺失导致牙釉质器官的多个细胞层发育不良，最终导致牙齿畸形和囊肿形成。具体而言，星状网状组织和中间层的细胞形成不良，导致囊性变化。前成釉细胞未能沿根尖轴延长，并持续强烈表达Sox2和P63，而在细胞分化过程中通常会下调Sox2和P63的表达。釉突标记的表达在突变体中也被减弱。随着时间的推移，突变体中的细胞增殖和细胞大小显着降低。重要的是，我们发现突变体中细胞骨架成分的数量减少和异常聚集，同时细胞骨架调节剂Cdc42的表达减弱，其上皮缺失导致相似​​的表型。此外，器官培养系统中肌动蛋白装配的破坏影响了牙胚的细胞增殖和细胞分化，支持了mTOR调控的细胞骨架动力学对mTOR突变小鼠表型的致病作用。为了进一步支持该观点，我们显示了mTOR过度激活导致牙釉质器官中细胞骨架成分的合成和组装增加，以及细胞分化加速。最后，我们证明了mTOR主要通过mTORC1途径调节釉质器官的发育。