GSE103349
Alteration of miRNome by recombinant ncRNA agents
Purpose: Identify whether recombinant ncRNAs are recognized by mammalian cells and processed into mature microRNAs, thus leading to the alteration of downstream target genes. In addition, we also explored whether cleavage is Dicer dependent. Methods: HEK293T (Dicer-WT) and 424 (Dicer-KO) cell lines were treated with 20nM of control tRNA (MSA), nCAR/miR-34a-5p, or nCAR/miR-34a-3p in triplicate. Results: Read counts of known miRNAs were analyzed using a miRDeep module, and read counts derived from nCAR/miR-34a-5p or nCAR/miR-124a-3p transfection were obtained by mapping sequence reads by an in-house developed PERL script. EdgeR was used to analyze significant (P-value < 0.05 and log2CPM > 6) differentially expressed miRNAs and sRNAs. Conclusions: Upon transfection of recombinant RNA agents in mammalian cells, nCAR/miR-34a-5p became the most dominant miRNA and was processed in a Dicer dependent manner, while nCAR/miR-124a-3p become the 7th most abundant miRNA and displayed comparable levels with or without Dicer. In both cases, different isoforms of the mature miRNA sequence were generated. Overall design: A total of 18 samples were analyzed. Three biological replicates in each group were subjected to treatment with control tRNA (MSA), nCAR/miR-34a-5p, or nCAR/miR-124a-3p. Therefore, each cell line had a total of 9 samples and the miRNome was explored in two different cell lines for a total of 18 samples.