Cell and sub-compartment specific proteomics reveals astrocyte contributions in a mouse model of OCD

We investigated the sub-proteomes of different functional compartments in astrocyte and neurons in the striatum. We compared the proteomes to the cell-specific RNA profiles of neurons and astrocytes from these analyses. We found one gene and protein that was highly expressed in neurons and astrocytes called SAPAP3. Knockout of this gene product results in an OCD-like phenotype in mouse. RNA-seq of astrocytes and bulk RNA-seq analyses of this mouse model revealed very low DEGs. 40 samples from 24 adult mice. Astrocyte RiboTag AAV and Neuron RiboTag AAV was injected into the striatum to extract RNA specifically from astrocytes or neurons. We used wildtype mice and used an OCD mouse model (SAPAP3 KO). After tissue homogenization, RNA was purified from the cleared lysate as the input sample and astrocyte or neuron specific ribosome-associated RNA was immunoprecipitated with a hemagglutinin (HA) tag as the IP sample.本研究针对纹状体中星形胶质细胞（astrocyte）与神经元内不同功能区室的亚蛋白质组展开探究。将上述得到的蛋白质组与本次分析中神经元及星形胶质细胞的细胞特异性RNA谱进行比对。本研究发现一个在神经元与星形胶质细胞中高表达的基因及对应蛋白质，命名为SAPAP3。该基因产物的敲除会使小鼠出现类强迫症（OCD）表型。对该小鼠模型的星形胶质细胞RNA测序（RNA-seq）及批量RNA测序分析显示，差异表达基因（DEGs）数量极少。实验共采集24只成年小鼠的40份样本。将星形胶质细胞核糖体标记腺相关病毒（RiboTag AAV）与神经元核糖体标记腺相关病毒注射至纹状体，以分别特异性提取星形胶质细胞或神经元内的RNA。本研究使用野生型小鼠及SAPAP3敲除（SAPAP3 KO）类强迫症小鼠模型。组织匀浆后，从澄清裂解液中纯化得到RNA作为输入样本；同时通过血凝素（HA）标签对星形胶质细胞或神经元特异性核糖体结合RNA进行免疫沉淀，得到免疫沉淀（IP）样本。