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Published on: July 29, 2016
Identification of the toxic 6mer seed consensus for human cancer cells
Monal Patel1, Elizabeth T Bartom2,3, Bidur Paudel4
1Department of Medicine/Division of Hematology/Oncology, Feinberg School of Medicine, Northwestern University, Chicago, IL, USA. monal.patel@northwestern.edu.
Abstract:
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 G-rich 6mers as the most toxic seeds. We have now tested two additional cell lines, one human and one mouse, identifying the GGGGGC consensus as the most toxic average 6mer seed for human cancer cells while slightly less significant for mouse cancer 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 located predominantly 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.
Insights
A novel cancer cell death mechanism, 6mer seed toxicity, utilizes specific RNA sequences to eliminate survival genes. G-rich sequences, particularly GGGGGC, show potent toxicity against human cancer cells, offering therapeutic potential.
Area of Science:
- Molecular Biology
- Cancer Research
- RNA Therapeutics
Background:
- 6mer seed toxicity is a novel mechanism for cancer cell death.
- This process involves the elimination of survival genes, termed Death Induced by Survival Gene Elimination (DISE).
- It relies on small interfering RNAs (siRNAs) or short hairpin RNAs (shRNAs) with specific G-rich nucleotide compositions.
Purpose of the Study:
- To identify the most toxic 6mer seeds for cancer cell death.
- To elucidate the mechanism by which the most toxic seed, GGGGGC, induces cancer cell death.
- To evaluate the therapeutic potential of siRNAs containing the GGGGGC seed.
Main Methods:
- Arrayed screening of 4096 6mer seeds across multiple human and mouse cell lines.
- RNA sequencing and bioinformatics analysis to identify gene targets of the GGGGGC seed.
- Functional assays to confirm direct targeting and the impact of seed match mutations.
Main Results:
- G-rich 6mer seeds were identified as the most toxic.
- The GGGGGC consensus sequence was the most toxic average 6mer seed for human cancer cells.
- siRNA containing the GGGGGC seed (siGGGGGC) targets GCCCCC seed matches in the 3' UTR of survival genes, inducing miRNA-like cell death.
Conclusions:
- The GGGGGC seed mediates cancer cell death through targeting critical survival genes.
- siGGGGGC exhibits miRNA-like activity, leading to cancer cell elimination.
- Artificial miRNAs, siRNAs, or shRNAs containing the GGGGGC seed represent a promising new avenue for cancer therapeutics.
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