Mutation in Wdr45 leads to early motor dysfunction and widespread aberrant axon terminals in a beta-propeller protein associated neurodegeneration (BPAN) patient-inspired mouse model

Beta-propeller Protein Associated Neurodegeneration (BPAN) is a devastating neurodevelopmental and neurodegenerative disease linked to variants in WDR45. Currently, there is no cure or disease altering treatment for this disease. This is, in part, due to a lack of insight into early phenotypes of BPAN progression and WDR45s role in establishing and maintaining neurological function. Here we generated and characterized a mouse model bearing a c52C>T BPAN patient variant in Wdr45. We show this mutation ablates WDR45 protein expression and alters autophagy in the brain. Behavioral analysis of these mice revealed characteristic signs of BPAN including cognitive impairment, hyperactivity, and motor decline. We show these behaviors coincide with widespread glial activation and development of axonal spheroids in multiple neuron subclasses throughout the brain. Several lines of evidence suggest these spheroids arise from axon terminals. Transcriptomic analysis uncovered multiple disrupted pathways in the cortex including genes associated with synapses, neurites, endosomes, endoplasmic reticulum, and ferroptosis. This is supported by accumulation of the iron regulating transferrin receptor 1 (TFRC) and the endoplasmic reticulum resident calreticulin (CALR) in the cortex as these animals age. CALR forms spheroid structures similar to the axonal spheroids seen in these animals. Taken together, our data suggest that WDR45 is necessary for healthy brain function and maintenance of axon terminals. This model opens the door to therapeutics targeting BPAN and further exploration of the role of WDR45 in neuronal function. Cortical samples from 3-month-old Wdr45 c52C>T mice and wildtype mice were collected and RNA was isolated using a TRIzol Chloroform-Isopropanol protocol. RNA sequencing experiments were then performed with analysis comparing transcriptomes between Wdr45 c52C>T mice and wildtype mice.β螺旋桨蛋白相关神经退行性疾病（Beta-propeller Protein Associated Neurodegeneration，BPAN）是一种与WDR45基因变异相关的致死性神经发育及神经退行性疾病，目前尚无治愈手段或可改变疾病病程的治疗方案。这在一定程度上源于对BPAN进展的早期表型以及WDR45在建立和维持神经系统功能中的作用缺乏深入认知。本研究构建并鉴定了一种携带Wdr45基因c52C>T BPAN患者致病变异的小鼠模型。研究显示，该突变可消除WDR45蛋白的表达，并改变大脑内的自噬（autophagy）过程。对该小鼠的行为学分析显示，其出现了BPAN的特征性表现：认知功能障碍、活动亢进与运动能力衰退。研究发现，此类行为异常与大脑内广泛的神经胶质激活以及多神经元亚群中轴突球体的形成同步发生。多项实验证据表明，此类轴突球体起源于轴突末梢。转录组学分析揭示，小鼠大脑皮层内多条通路发生紊乱，涉及突触、神经突、内体、内质网及铁死亡（ferroptosis）相关基因。随着小鼠年龄增长，皮层内铁调节蛋白转铁蛋白受体1（transferrin receptor 1，TFRC）与内质网驻留蛋白钙网蛋白（calreticulin，CALR）的积累进一步验证了上述结论。CALR可形成与该小鼠体内所见轴突球体结构相似的球体样聚集体。综上，本研究数据表明，WDR45对于维持大脑正常功能及轴突末梢的稳态至关重要。该小鼠模型为BPAN的靶向治疗研究提供了重要契机，同时也为进一步探索WDR45在神经元功能中的作用开辟了新的研究方向。本研究收集了3月龄Wdr45 c52C>T突变小鼠与野生型小鼠的皮层组织，采用TRIzol-氯仿-异戊醇法提取总RNA，随后通过RNA测序（RNA sequencing）技术比较两组小鼠的皮层转录组差异。