Effect of Ephrin-A5 DNA Nanocalipers stimulation of EphA2 receptor signaling in U3013 cell line

Ligand binding induces extensive spatial reorganization and clustering of the EphA2 receptor at the cell membrane. It has previously been shown that the nanoscale spatial distribution of ligands modulates EphA2 receptor reorganization, activation and the invasive properties of cancer cells. However, the mechanisms by which cells transduce ligand nanoscale spatial distribution signals have not been elucidated. Here we used DNA origami nanocalipers to control the positions of ephrin-A5 ligands at the nanoscale and investigated the transcriptional responses following ligand binding. Using mRNA sequencing, we determined the transcriptional profiles of glioblastoma cells treated with nanocalipers presenting a single ephrin-A5 dimer or two dimers spaced 14, 40 or 100 nm apart. We observed divergent transcriptional responses to ephrin-A5 nano-organization, with ephrin-A5 dimers spaced 40 or 100 nm apart showing the highest levels of differential expressed genes compared to treatment with n anocalipers that do not present ephrin-A5. These findings show that the nanoscale organization of ephrin-A5 modulates transcriptional responses to EphA2 activation. Here, we compare differently spaced Ephrin-A5 dimers on DNA nanocalipers and the effect they have on the U3013 transcriptome when stimulating the EphA2 receptor