ScRNA-seq of Brca2 mammary organoids reveals a subpopulation expansion in response to replication stress

BRCA2 mutation carriers preferentially develop luminal-like breast cancers, but it remains unclear how BRCA2 mutations affect mammary epithelial subpopulations. Here, we report that Brca2mut/WT mammary organoids subjected to replication stress activated a transcriptional response that selectively expands Brca2mut/WT luminal cells lacking hormone receptor expression (HR-). While CyTOF analyses revealed comparable epithelial compositions among wildtype and Brca2mut/WT mammary glands, Brca2mut/WT HR- luminal cells exhibited greater organoid formation and preferentially survived and expanded under replication stress. ScRNA-seq analysis corroborated the expansion of HR- luminal cells which express elevated levels of Tetraspanin-8 (Tspan8) and Thrsp mRNA, and pathways implicated in replication stress survival including Type I interferon responses. Notably, CRISPR/Cas9-mediated deletion of Tspan8 or Thrsp prevented Brca2mut/WT HR- luminal cell expansion. Our findings indicate that Brca2mut/WT cells have an activate a transcriptional response after replication stress that preferentially favours outgrowth of HR- luminal cells through the expression of interferon-responsive and mammary alveolar genes. In this study we utilised a genetically engineered murine model carrying a monoallelic germline Brca2 truncating mutation, Brca2mut/WT. organoids were generated from Brca2mut/WT and wildtype mammary glands, which were exposed to replication stresses at regular intervals over a two month period. We performed scRNA-seq from early (passage 0) and late (passage 4) organoid culture time points. The data provides xx scRNA-seq profiles using 10X Genomics Chromium platform, compromising a total of 34,200 from 3 independent experiments. This includes the four different conditions (wildtype or Brca2mut/WT with or without HU treatments) and time points (Passage 0 or 4).