Identification of the toxic 6mer seed consensus for human cancer cells

6mer seed toxicity is a novel cell death mechanism that kills cancer cells by triggering death induced by survival gene elimination (DISE). It is based on si- or shRNAs with a specific G-rich nucleotide composition in position 2-7 of their guide strand. An arrayed screen of 4096 6mer seeds on two human and two mouse cell lines identified a consensus GGGGGC as the most toxic seed. We have now tested two additional cell lines, one human and one mouse, confirming the GGGGGC consensus as the most toxic average 6mer seed for human cells while slightly less significant for mouse cells. RNA Seq and bioinformatics analyses suggested that an siRNA containing the GGGGGC seed (siGGGGGC) is toxic to cancer cells by targeting GCCCCC seed matches in the 3' UTR of a set of genes critical for cell survival. We have identified several genes targeted by this seed and demonstrate direct and specific targeting of GCCCCC seed matches, which is attenuated upon mutation of the GCCCCC seed matches in these 3' UTRs. Our data show that siGGGGGC kills cancer cells through its miRNA-like activity and points at artificial miRNAs, si- or shRNAs containing this seed as a potential new cancer therapeutics PC3 or HeyA8 cells were transfected with different siRNAs at 1nM and 24 or 48hrs after transfection RNA was isolated and subjetced to lareg RNA Seq analysis