PT  - JOURNAL ARTICLE
AU  - Yujie Zhang
AU  - Vicent Pelechano
TI  - High-throughput 5’P sequencing reveals environmental regulated ribosome stalls at termination level
AID  - 10.1101/2020.06.22.165134
DP  - 2020 Jan 01
TA  - bioRxiv
PG  - 2020.06.22.165134
4099  - http://biorxiv.org/content/early/2020/06/23/2020.06.22.165134.short
4100  - http://biorxiv.org/content/early/2020/06/23/2020.06.22.165134.full
AB  - RNA degradation is critical for gene expression and mRNA quality control. mRNA degradation is intimately connected to the translation process up to the degree that 5’-3’ co-translational mRNA degradation can be used as a proxy for ribosome dynamics. Here we present an improved high-throughput 5’P RNA sequencing method (HT-5Pseq). HT-5Pseq is easy, scalable and uses affordable duplex-specific nuclease based rRNA depletion. We investigate in vivo ribosome stalls in Saccharomyces cerevisiae and Schizosaccharomyces pombe focusing on translation termination. We show that degradation intermediates of mRNAs with high termination pausing often use TGA as stop codon preceded by Lys and have relatively shorter poly(A) tails. We investigate the regulation of this process after glucose deprivation and reveal that, in addition to translation initiation, glucose starvation also leads to ribosome stall at the stop codon. Finally, we investigate gene-specific termination pausing across multiple conditions and show that it is a highly regulated process that crosstalks with codon optimality and the length of poly(A) remaining after deadenylation. In summary, HT-5Pseq is an improved and cost-effective approach that allows to investigate the crosstalk between RNA degradation and the translation process and that is particularly useful to examine its coupling at the level of translation termination.Key pointsHT-5Pseq facilitates high throughput investigation of 5’P mRNA degradation intermediates.Gene-specific ribosome pause at termination level is common and environmentally regulated.Ribosome stall at termination level are associated with TGA usage and shorter deadenylated poly(A) tails.Competing Interest StatementThe authors have declared no competing interest.