p53 governs an alveolar type 1 differentiation program in lung cancer suppression [scRNA-Seq]

Lung cancer is the leading cause of cancer deaths worldwide. TP53 tumor suppressor gene mutations occur in 50% of lung adenocarcinomas (LUADs) and are linked to poor prognosis, but how p53 suppresses LUAD development remains enigmatic. We show here that p53 suppresses LUAD by governing cell state, by promoting alveolar type 1 (AT1) differentiation. Using mice expressing oncogenic Kras and null, wild-type, or hypermorphic p53 alleles in alveolar type 2 (AT2) cells, we observed graded effects of p53 on LUAD initiation and progression. RNA-sequencing and ATAC-sequencing of LUAD cells uncovered a p53-induced AT1 differentiation program during tumor suppression in vivo through direct DNA binding, chromatin remodeling, and AT1 gene induction. Single-cell transcriptomics analyses revealed that during LUAD evolution, p53 promotes AT1 differentiation through action in a transitional cell state analogous to a transient intermediary seen during AT2-to-AT1 differentiation in alveolar injury repair. Notably, p53 inactivation resulted in the inappropriate persistence of these transitional cancer cells accompanied by upregulated growth signaling and divergence from lung lineage identity, characteristics associated with LUAD progression. Analysis of p53wt and p53-null mice showed that p53 also directs alveolar regeneration after injury, by regulating AT2 self-renewal and promoting transitional cell differentiation into AT1 cells. Collectively, these findings illuminate mechanisms of p53-mediated LUAD suppression, in which p53 governs alveolar differentiation, and suggest that tumor suppression reflects a fundamental role of p53 in orchestrating tissue repair after injury. Mouse lung cells during injury were isolated via FACS and analyzed via 10x scRNA-seq

肺癌是全球范围内癌症相关死亡的首要诱因。TP53肿瘤抑制基因（TP53 tumor suppressor gene）突变在50%的肺腺癌（LUAD）中发生，且与不良预后密切相关，但p53抑制肺腺癌发生发展的具体机制仍未阐明。本研究证实，p53可通过调控细胞状态、促进肺泡1型（AT1）细胞分化来发挥肺腺癌抑制作用。我们构建了在肺泡2型（AT2）细胞中表达致癌性Kras以及缺失型、野生型或功能增强型p53等位基因的小鼠模型，借此观察到p53对肺腺癌的起始与进展存在梯度调控效应。通过对肺腺癌细胞进行RNA测序（RNA-seq）与转座酶可及性染色质测序（ATAC-seq），我们发现，在体内肿瘤抑制过程中，p53可通过直接结合DNA、重塑染色质并诱导AT1相关基因表达，激活一套AT1分化程序。单细胞转录组学分析显示，在肺腺癌演化进程中，p53可通过作用于一类过渡细胞状态促进AT1分化，该状态与肺泡损伤修复过程中AT2细胞向AT1细胞分化时出现的瞬时中间态类似。值得注意的是，p53失活会导致这类过渡性癌细胞异常持续存在，同时伴随生长信号上调与肺谱系身份的偏离，这些特征均与肺腺癌进展密切相关。对野生型p53与p53敲除小鼠的分析表明，p53还可通过调控AT2细胞自我更新、促进过渡细胞分化为AT1细胞，从而介导损伤后的肺泡再生。综上，本研究阐明了p53介导的肺腺癌抑制机制——即p53调控肺泡分化进程，并提示肿瘤抑制本质上反映了p53在协调损伤后组织修复过程中的核心功能。本研究通过荧光激活细胞分选术（FACS）分离损伤后的小鼠肺细胞，并采用10×单细胞RNA测序（10x scRNA-seq）完成相关分析。