RT Journal Article SR Electronic T1 ContactBlot: Microfluidic Control and Measurement of Cell-Cell Contact State to Assess Contact-Inhibited ERK Signaling JF bioRxiv FD Cold Spring Harbor Laboratory SP 2023.11.06.565857 DO 10.1101/2023.11.06.565857 A1 Zhang, Yizhe A1 Naguro, Isao A1 Ryuno, Hiroki A1 Herr, Amy E. YR 2024 UL http://biorxiv.org/content/early/2024/06/06/2023.11.06.565857.1.abstract AB Extracellular signal-regulated kinase (ERK) signaling is essential to regulated cell behaviors, including cell proliferation, differentiation, and apoptosis. The influence of cell-cell contacts on ERK signaling is central to epithelial cells, yet few studies have sought to understand the same in cancer cells, particularly with single-cell resolution. To acquire same-cell measurements of both phenotypic (cell-contact state) and targeted-protein profile (ERK phosphorylation), we prepend high-content, whole-cell imaging prior to endpoint cellular-resolution western blot analyses for each of hundreds of individual HeLa cancer cells cultured on that same chip, which we call contactBlot. By indexing the phosphorylation level of ERK in each cell or cell-cluster to the imaged cell-contact state, we compare ERK signaling between isolated and in-contact cells. We observe attenuated (∼2×) ERK signaling in HeLa cells which are in-contact versus isolated. Attenuation is sustained when the HeLa cells are challenged with hyperosmotic stress. Our findings show the impact of cell-cell contacts on ERK activation with isolated and in-contact cells, while introducing a multi omics tool for control and scrutiny of cell-cell interactions.Competing Interest StatementThe authors have declared no competing interest.