Abstract：
(1) Background: Activity-dependent neuroprotective protein (ADNP) is essential for neuronal structure and function. Multiple de novo pathological mutations in ADNP cause the autistic ADNP syndrome, and they have been further suggested to affect Alzheimer\'s disease progression in a somatic form. Here, we asked if different ADNP mutations produce specific neuronal-like phenotypes toward better understanding and personalized medicine. (2) Methods: We employed CRISPR/Cas9 genome editing in N1E-115 neuroblastoma cells to form neuron-like cell lines expressing ADNP mutant proteins conjugated to GFP. These new cell lines were characterized by quantitative morphology, immunocytochemistry and live cell imaging. (3) Results: Our novel cell lines, constitutively expressing GFP-ADNP p.Pro403 (p.Ser404* human orthologue) and GFP-ADNP p.Tyr718* (p.Tyr719* human orthologue), revealed new and distinct phenotypes. Increased neurite numbers (day 1, in culture) and increased neurite lengths upon differentiation (day 7, in culture) were linked with p.Pro403*. In contrast, p.Tyr718* decreased cell numbers (day 1). These discrete phenotypes were associated with an increased expression of both mutant proteins in the cytoplasm. Reduced nuclear/cytoplasmic boundaries were observed in the p.Tyr718* ADNP-mutant line, with this malformation being corrected by the ADNP-derived fragment drug candidate NAP. (4) Conclusions: Distinct impairments characterize different ADNP mutants and reveal aberrant cytoplasmic-nuclear crosstalk.
摘要：
(1)研究背景:活动依赖性神经保护蛋白(ADNP)对神经元的结构和功能至关重要。ADNP中的多个从头病理突变导致自闭症ADNP综合征，并进一步建议它们以躯体形式影响阿尔茨海默病的进展。这里,我们询问不同的ADNP突变是否会产生特定的神经元样表型,以更好地理解和个性化医疗.（2）方法：我们在N1E-115神经母细胞瘤细胞中使用CRISPR/Cas9基因组编辑以形成表达与GFP缀合的ADNP突变蛋白的神经元样细胞系。这些新细胞系的特征在于定量形态学,免疫细胞化学和活细胞成像。（3）结果：我们的新细胞系，组成型表达GFP-ADNPp.Pro403（p.Ser404*人类直向同源物）和GFP-ADNPp.Tyr718*（p。Tyr719*人类直系同源），揭示了新的和独特的表型。增加的神经突数量(第1天,在培养中)和分化后增加的神经突长度(第7天,在培养中)与p.Pro403*相关。相比之下,p.Tyr718*减少细胞数(第1天)。这些离散的表型与细胞质中两种突变蛋白的表达增加有关。在p.Tyr718*ADNP突变体系中观察到核/细胞质边界减少，这种畸形被ADNP衍生的片段药物候选NAP纠正。（4）结论：不同的ADNP突变体具有明显的损伤特征，并揭示了异常的细胞质-核串扰。
