Genomic Medicine Institute, Lerner Research Institute, Cleveland Clinic, Cleveland, OH, 44195, USA.
Cleveland Clinic Lerner College of Medicine, Case Western Reserve University, 9500 Euclid Avenue, Cleveland, OH, 44195, USA.
Mol Autism. 2020 Jun 1;11(1):43. doi: 10.1186/s13229-020-00337-2.
PTEN, a syndromic autism spectrum disorder (ASD) risk gene, is mutated in approximately 10% of macrocephalic ASD cases. Despite the described genetic association between PTEN and ASD and ensuing studies, we continue to have a limited understanding of how PTEN disruption drives ASD pathogenesis and maintenance.
We derived neural stem cells (NSCs) from the dentate gyrus (DG) of Pten mice, a model that recapitulates PTEN-ASD phenotypes. We subsequently characterized the expression of stemness factors, proliferation, and differentiation of neurons and glia in Pten NSCs using immunofluorescent and immunoblotting approaches. We also measured Creb phosphorylation by Western blot analysis and expression of Creb-regulated genes with qRT-PCR.
The m3m4 mutation decreases Pten localization to the nucleus and its global expression over time. Pten NSCs exhibit persistent stemness characteristics associated with increased proliferation and a resistance to neuronal maturation during differentiation. Given the increased proliferation of Pten NSCs, a significant increase in the population of immature neurons relative to mature neurons occurs, an approximately tenfold decrease in the ratio between the homozygous mutant and wildtype. There is an opposite pattern of differentiation in some Pten glia, specifically an increase in astrocytes. These aberrant differentiation patterns associate with changes in Creb activation in Pten NSCs. We specifically observed loss of Creb phosphorylation at S133 in Pten NSCs and a subsequent decrease in expression of Creb-regulated genes important to neuronal function (i.e., Bdnf). Interestingly, Bdnf treatment is able to partially rescue the stunted neuronal maturation phenotype in Pten NSCs.
Constitutional disruption of Pten nuclear localization with subsequent global decrease in Pten expression generates abnormal patterns of differentiation, a stunting of neuronal maturation. The propensity of Pten disruption to restrain neurons to a more progenitor-like state may be an important feature contributing to PTEN-ASD pathogenesis.
PTEN 是一种综合征性自闭症谱系障碍（ASD）风险基因，约 10%的大头 ASD 病例存在 PTEN 突变。尽管已经描述了 PTEN 与 ASD 之间的遗传关联以及随后的研究，但我们对 PTEN 缺失如何驱动 ASD 发病机制和维持的了解仍然有限。
我们从 Pten 小鼠的齿状回（DG）中衍生出神经干细胞（NSC），这是一种重现 PTEN-ASD 表型的模型。随后，我们使用免疫荧光和免疫印迹方法来表征 Pten NSCs 中的干性因子表达、增殖和神经元及神经胶质分化。我们还通过 Western blot 分析测量了 Creb 的磷酸化水平，并通过 qRT-PCR 检测了 Creb 调节基因的表达。
m3m4 突变会减少 Pten 向核内的定位及其随时间推移的整体表达。Pten NSCs 表现出持续的干性特征，与增殖增加和分化过程中神经元成熟的抵抗力有关。由于 Pten NSCs 的增殖增加，未成熟神经元的群体相对于成熟神经元显著增加，杂合突变体与野生型之间的比例约减少十倍。在一些 Pten 神经胶质中出现相反的分化模式，特别是星形胶质细胞增加。这些异常分化模式与 Pten NSCs 中 Creb 激活的变化有关。我们特别观察到 Pten NSCs 中 Creb 的 S133 磷酸化丧失，以及对神经元功能重要的 Creb 调节基因的表达随后下降（即 Bdnf）。有趣的是，BDNF 处理能够部分挽救 Pten NSCs 中神经元成熟受阻的表型。
PTEN 核定位的结构破坏，随后 PTEN 表达的整体减少，会产生异常的分化模式，阻止神经元成熟。PTEN 缺失抑制神经元向更祖细胞样状态的倾向可能是导致 PTEN-ASD 发病机制的一个重要特征。
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