Journal: Nature Communications

Article Title: BdLT-Seq as a barcode decay-based method to unravel lineage-linked transcriptome plasticity

doi: 10.1038/s41467-023-36744-1

Figure Lengend Snippet: a Scheme representing the workflow to generate populations of cells resistant to TRAIL-induced apoptosis (either induced in response to or selected for) and its processing through the BdLT-Seq to explore transcriptome divergence upon TRAIL withdrawal. b Histogram depicting TRAIL-induced apoptosis in HA1ER parental clone (F12, pink) and two F12-subclones (1F8 and 1C9, blue and light purple respectively) as determined by cleaved PARP staining analysed by flow cytometry. No TRAIL (NT) and constant rhTRAIL treatment are depicted as controls of acquired TRAIL resistance. Day 1 to Day 3 of rhTRAIL withdrawal is shown to denote escape from TRAIL resistance and reversion to fractional killing induced by TRAIL. Histograms show the mean value ± standard deviation (SD) of three independent biological replicates (~20,000 cells per condition). Statistical significance was assessed by a two-tailed paired Student’s t -test. *** P value < 0.0005, ** P value < 0.005, * P value < 0.05. The clonal relationship is also depicted. Individual data points are shown. c UMAP plots of scRNA-Seq data showing transcriptome states identified for F12, 1F8 and 1C9 cells at Day 0 (TRAIL-resistant, left panel) and Day 3 (TRAIL withdrawal, right panel) of tracing. Gene expression state boundaries are depicted as coloured lattices. The clonal relationship is also depicted. d Chord diagrams depicting transcriptome state dynamics for F12, 1F8 and 1C9 cells that belong to a particular state/cluster at Day 0 of tracing (TRAIL-resistant) and their divergence after 3 days (TRAIL withdrawal). All detected clusters are depicted (C0 to C10) and integrate the collapsed behaviour of all cells that belong to each particular gene expression state. The origin cluster is depicted in pink (Day 0) and chords represent endpoint cluster association (Day 3). Source data are provided as a Source Data file.

Article Snippet: Immunolabelling of cleaved PARP was performed using fluorescently labelled Cleaved PARP Alexa 647 antibody (Cell Signalling, cat. #6987) (1:200 dilution) for 1 h at 20 °C.

Techniques: Staining, Flow Cytometry, Standard Deviation, Two Tailed Test, Expressing