Metastasis-initiating cells dynamically adapt to the distinct microenvironments of different organs, but these early adaptations are poorly understood due to the limited sensitivity of in situ transcriptomics. Researchers from Memorial Sloan-Kettering Cancer Center developed fluorouracil-labeled RNA sequencing (Flura-seq) for in situ analysis with high sensitivity. Flura-seq utilizes cytosine deaminase (CD) to convert fluorocytosine to fluorouracil, metabolically labeling nascent RNA in rare cell populations in situ for purification and sequencing. Flura-seq revealed hundreds of unique, dynamic organ-specific gene signatures depending on the microenvironment in mouse xenograft breast cancer micrometastases. Specifically, the mitochondrial electron transport Complex I, oxidative stress and counteracting antioxidant programs were induced in pulmonary micrometastases, compared to mammary tumors or brain micrometastases. The researchers confirmed lung metastasis-specific increase in oxidative stress and upregulation of antioxidants in clinical samples, thus validating Flura-seq’s utility in identifying clinically actionable microenvironmental adaptations in early metastasis. The sensitivity, robustness and economy of Flura-seq are broadly applicable beyond cancer research.
Cell-type specific labeling and isolation of RNAs by Flura-tagging
(A) Schematic diagram showing RNA labeling and isolation using CD and 5-FC; (B) Chemical reactions steps involved in the labeling of RNA using CD and 5-FC