BioProject

mRNA endogenous decay profiles approach in 293T, HeLa and RPE cells. mRNA translation decodes nucleotide into amino acid sequences. More...

mRNA endogenous decay profiles approach in 293T, HeLa and RPE cells. mRNA translation decodes nucleotide into amino acid sequences. However, translation has also been shown to affect mRNA stability depending on codon composition in model organisms, although universality of this mechanism remains unclear. Here, using three independent approaches to measure exogenous and endogenous mRNA decay, we define which codons are associated with stable or unstable mRNAs in human cells. We demonstrate that the regulatory information affecting mRNA stability is encoded in codons and not in nucleotides. Stabilizing codons tend to be associated with higher tRNA levels and higher charged/total tRNA ratios. While mRNAs enriched in destabilizing codons tend to possess shorter poly(A)-tails, the poly(A)-tail is not required for the codon-mediated mRNA stability. This mechanism depends on translation; however, the number of ribosome loads into a mRNA modulates the codon-mediated effects on gene expression. This work provides definitive evidence that translation strongly affects mRNA stability in a codon-dependent manner in human cells. Overall design: 3 different cells with different timepoint, Cells were sub-cultured in 24 well plate overnight, so the cells were at around 80% coverage before treatment. Actinomycin D was added into the well, with final concentration of (5ug/ml), in 0.1% DMSO. Cells were directly collected with Trizol, at desired time-point for RNA extract. RNA-seq with ribo depletion was done for each sample, to calculate mRNA decay based on time-course mRNA-seq. For 293t, it is also treated with Cycloheximide. It was added at 2ug/ml, in H2O. "ad" in sample name means actinomycin D treatment, "chx" in the name means Cycloheximide. Less...