Biology subjects
Publications and source records attributed to Chyzynska, K..
Withdrawal StatementThe authors have withdrawn their manuscript owing to not all authors having approved the manuscript. Therefore, the authors do not wish this work to be cited as reference for the project. If you have any questions, please contact the corresponding author.
The rate of translation can vary considerably depending on the mRNA template. During the elongation phase the ribosome can transiently pause or permanently stall. A pause can provide the nascent protein with the required time to fold or be transported, while stalling can serve as quality control and trigger degradation of aberrant mRNA and peptide. Ribosome profiling has allowed for the genome-wide detection of such pause and stall sites, but due to library-specific biases, these predictions are often unreliable. Here, we address this by taking advantage of the deep conservation of the protein synthesis machinery, hypothesizing that similar conservation could exist for functionally important positions of ribosome slowdown - here collectively called stall sites. We analyze multiple ribosome profiling datasets from a phylogenetically diverse group of eukaryotes: yeast, fruit fly, zebrafish, mouse, and human and identify conserved stall sites. We find thousands of stall sites across multiple species, with proline, glycine, and negatively charged amino acids being the main facilitators of stalling. Many of the sites are found in RNA processing genes, suggesting that stalling might have a conserved regulatory effect on RNA metabolism. In summary, our results provide a rich resource for the study of conserved stalling and indicate possible roles of stalling in gene regulation.
The emergence of ribosome profiling to map actively translating ribosomes has laid the foundation for a diverse range of studies on translational regulation. The data obtained with different variations of this assay is typically manually processed, which has created a need for tools that would streamline and standardize processing steps.\n\nWe present Shoelaces, a toolkit for ribosome profiling experiments automating read selection and filtering to obtain genuine translating footprints. Based on periodicity, favoring enrichment over the coding regions, it determines the read lengths corresponding to bona fide ribosome protected fragments. The specific codon under translation (P-site) is determined by automatic offset calculations resulting in sub-codon resolution. Shoelaces provides both a user-friendly graphical interface for interactive visualisation in a genome browser-like fashion and a command line interface for integration into automated pipelines. We process 79 libraries and show that studies typically discard excessive amounts of data in their manual analysis pipelines.\n\nShoelaces streamlines ribosome profiling analysis offering automation of the processing, a range of interactive visualization features and export of the data into standard formats. Shoelaces stores all processing steps performed in an XML file that can be used by other groups to exactly reproduce the processing of a given study. We therefore anticipate that Shoelaces can aid researchers by automating what is typically performed manually and contribute to the overall reproducibility of studies. The tool is freely distributed as a Python package, with additional instructions and demo datasets available at https://bitbucket.org/valenlab/shoelaces