Metastable Intermediates Identified in Epithelial to Mesenchymal Transition are Regulated by G-quadruplex DNA Structures

Cancer is a heterogenous disease, with multiple cellular subpopulations present within a single tumour mass that differ genetically and morphologically, and thus respond differently to chemotherapeutics. Epithelial-to-Mesenchymal transition (EMT) has been shown to play a role in tumour heterogeneity. Single-cell sequencing is critical to identify cell-type-specific transcriptomic differences with multiplexing methods increasing experimental scope with reduced cost. Cell hashing with barcoded antibodies is commonly used to multiplex samples but is limited to samples expressing target antigens. Antigen-independent methods of barcoding cells, such as barcoded lipid-anchors, have gained traction but present substantial populations that cannot be unambiguously demultiplexed. Herein we report a multiplexed single-cell transfection-enabled cell hashing sequencing (scTECH-seq) platform, which uses antigen-independent endocytic uptake to barcode cells, resulting in efficient, uniform barcoding with high cell recovery. We apply this methodology to identify distinct metastable cell states in human mammary cells undergoing EMT and show that stabilisation of G-quadruplex DNA has the potential to inhibit EMT. We performed single-cell RNA-seq of human mammary epithelial cells to identify metastable intermediate cell states in TGF-β induced epithelial-to-mesenchymal transition (EMT). We also treated cells with pyridostatin to investigate the role G-quadruplex structures play in EMT. We combined this with our new method scTECH-seq to multiplex samples and quantify transcripts in the presence and absence of pyridostatin in TGF-β induced EMT. Please note that processed data generated from both [GEX] and [CBO] samples, is linked to the corresonding [GEX] sample records.