Dataset Information

High-Resolution Ribosome Profiling Defines Discrete Ribosome Elongation States and Translational Regulation during Cellular Stress.

ABSTRACT: Ribosomes undergo substantial conformational changes during translation elongation to accommodate incoming aminoacyl-tRNAs and translocate along the mRNA template. We used multiple elongation inhibitors and chemical probing to define ribosome conformational states corresponding to differently sized ribosome-protected mRNA fragments (RPFs) generated by ribosome profiling. We show, using various genetic and environmental perturbations, that short 20-22 or classical 27-29 nucleotide RPFs correspond to ribosomes with open or occupied ribosomal A sites, respectively. These distinct states of translation elongation are readily discerned by ribosome profiling in all eukaryotes we tested, including fungi, worms, and mammals. This high-resolution ribosome profiling approach reveals mechanisms of translation-elongation arrest during distinct stress conditions. Hyperosmotic stress inhibits translocation through Rck2-dependent eEF2 phosphorylation, whereas oxidative stress traps ribosomes in a pre-translocation state, independent of Rck2-driven eEF2 phosphorylation. These results provide insights and approaches for defining the molecular events that impact translation elongation throughout biology.

SUBMITTER: Wu CC

PROVIDER: S-EPMC6411040 | biostudies-literature | 2019 Mar

REPOSITORIES: biostudies-literature

ACCESS DATA

Publications

High-Resolution Ribosome Profiling Defines Discrete Ribosome Elongation States and Translational Regulation during Cellular Stress.

Wu Colin Chih-Chien CC Zinshteyn Boris B Wehner Karen A KA Green Rachel R

Molecular cell 20190124 5

Ribosomes undergo substantial conformational changes during translation elongation to accommodate incoming aminoacyl-tRNAs and translocate along the mRNA template. We used multiple elongation inhibitors and chemical probing to define ribosome conformational states corresponding to differently sized ribosome-protected mRNA fragments (RPFs) generated by ribosome profiling. We show, using various genetic and environmental perturbations, that short 20-22 or classical 27-29 nucleotide RPFs correspond ...[more]

PMID: 30686592

Similar Datasets

Project description:Vaccinia virus infection causes a host shutoff that is marked by global inhibition of host protein synthesis. Though the host shutoff may facilitate reallocation of cellular resources for viral replication and evasion of host antiviral immune responses, it poses a challenge for continuous synthesis of cellular proteins that are important for viral replication. It is, however, unclear whether and how certain cellular proteins may be selectively synthesized during the vaccinia virus-induced host shutoff. Using simultaneous RNA sequencing and ribosome profiling, two techniques quantifying genome-wide levels of mRNA and active protein translation, respectively, we analyzed the responses of host cells to vaccinia virus infection at both the transcriptional and translational levels. The analyses showed that cellular mRNA depletion played a dominant role in the shutoff of host protein synthesis. Though the cellular mRNAs were significantly reduced, the relative translation efficiency of a subset of cellular mRNAs increased, particularly those involved in oxidative phosphorylation that are responsible for cellular energy production. Further experiments demonstrated that the protein levels and activities of oxidative phosphorylation increased during vaccinia virus infection, while inhibition of the cellular oxidative phosphorylation function significantly suppressed vaccinia virus replication. Moreover, the short 5' untranslated region of the oxidative phosphorylation mRNAs contributed to the translational upregulation. These results provide evidence of a mechanism that couples translational control and energy metabolism, two processes that all viruses depend on host cells to provide, to support vaccinia virus replication during a host shutoff.IMPORTANCE Many viral infections cause global host protein synthesis shutoff. While host protein synthesis shutoff benefits the virus by relocating cellular resources to viral replication, it also poses a challenge to the maintenance of cellular functions necessary for viral replication if continuous protein synthesis is required. Here we measured the host mRNA translation rate during a vaccinia virus-induced host shutoff by analyzing total and actively translating mRNAs in a genome-wide manner. This study revealed that oxidative phosphorylation mRNAs were translationally upregulated during vaccinia virus-induced host protein synthesis shutoff. Oxidative phosphorylation is the major cellular energy-producing pathway, and we further showed that maintenance of its function is important for vaccinia virus replication. This study highlights the fact that vaccinia virus infection can enhance cellular energy production through translational upregulation in the context of an overall host protein synthesis shutoff to meet energy expenditure.

Dataset Information

High-Resolution Ribosome Profiling Defines Discrete Ribosome Elongation States and Translational Regulation during Cellular Stress.

Publications

High-Resolution Ribosome Profiling Defines Discrete Ribosome Elongation States and Translational Regulation during Cellular Stress.

Similar Datasets

OmicsDI is part of the ELIXIR infrastructure

Tweets