Department of Pediatric Oncology, Dana-Farber Cancer Institute and Harvard Medical School, Boston, MA, USA; Broad Institute of MIT and Harvard, Cambridge, MA, USA; Chemical Biology Program, Harvard University, Cambridge, MA, USA.
Broad Institute of MIT and Harvard, Cambridge, MA, USA; Department of Biology, Massachusetts Institute of Technology, Cambridge, MA, USA.
Mol Cell. 2023 Apr 20;83(8):1350-1367.e7. doi: 10.1016/j.molcel.2023.03.013. Epub 2023 Apr 6.
The mammalian SWI/SNF (mSWI/SNF or BAF) family of chromatin remodeling complexes play critical roles in regulating DNA accessibility and gene expression. The three final-form subcomplexes-cBAF, PBAF, and ncBAF-are distinct in biochemical componentry, chromatin targeting, and roles in disease; however, the contributions of their constituent subunits to gene expression remain incompletely defined. Here, we performed Perturb-seq-based CRISPR-Cas9 knockout screens targeting mSWI/SNF subunits individually and in select combinations, followed by single-cell RNA-seq and SHARE-seq. We uncovered complex-, module-, and subunit-specific contributions to distinct regulatory networks and defined paralog subunit relationships and shifted subcomplex functions upon perturbations. Synergistic, intra-complex genetic interactions between subunits reveal functional redundancy and modularity. Importantly, single-cell subunit perturbation signatures mapped across bulk primary human tumor expression profiles both mirror and predict cBAF loss-of-function status in cancer. Our findings highlight the utility of Perturb-seq to dissect disease-relevant gene regulatory impacts of heterogeneous, multi-component master regulatory complexes.
哺乳动物 SWI/SNF（mSWI/SNF 或 BAF）家族的染色质重塑复合物在调节 DNA 可及性和基因表达方面发挥着关键作用。三个最终形式的亚复合物-cBAF、PBAF 和 ncBAF-在生化成分、染色质靶向以及在疾病中的作用方面存在明显差异；然而，其组成亚基对基因表达的贡献仍未完全确定。在这里，我们针对 mSWI/SNF 亚基个体和特定组合进行了基于 Perturb-seq 的 CRISPR-Cas9 敲除筛选，随后进行了单细胞 RNA-seq 和 SHARE-seq。我们发现了复杂、模块和亚基特异性对不同调控网络的贡献，并定义了旁系亚基关系，并在扰动时改变了亚复合物的功能。亚基之间的协同、内复合物遗传相互作用揭示了功能冗余和模块性。重要的是，单细胞亚基扰动特征映射到批量原发性人类肿瘤表达谱上，既反映又预测了癌症中 cBAF 功能丧失状态。我们的研究结果强调了 Perturb-seq 用于剖析异质、多成分主调控复合物与疾病相关基因调控影响的实用性。
