effect of RNA uptake on gene expressions in human NK and CD8-positive T cells

In the blood, non-vesicular extracellular RNA functions as a cell-cell communication molecule. We have identified that IL1b-mRNA fragment binds ZC3H12D protein on the cell surface. This protein expresses on NK and T cells. When extracellular RNA binds ZC3H12D on the cell surface, the RNA-ZC3H12D complex goes inside the cell to modulate cell character, and as a result, the RNA stimulated-immune cell became anti-tumor rather than pro-tumor. To decipher the molecular mechanisms to govern the extracellular RNA-mediated signaling, we carried out NGS analysis to find out the difference in gene expression profiles after external RNA stimulation. Human PBMC was purchased from Roche. Thawed PBMC was maintained in HPLM (Thermo) medium supplemented with 10% FCS. The following day, FITC labeled IL1b-mRNA (10ng/mL) was applied in the media, and 0.5h or 6h later cells were FACS-sorted to isolate NK cells (CD56+CD3-) and CD8+T cells. Isolated cells were immediately placed into Trizol (Thermo) reagent to obtain total RNA.