1. Development of GWIPS-viz genome browser Christof De Bo LAPTI, Biochemistry Department, University College Cork, Ireland

2. THEORY Ribosome profiling Introduction Ribosome profiling technique Initiating and elongating ribosomes GWIPS-viz Introduction Workflow Coverage plots vs. profiles Visualizations CONCLUSION INTRODUCTION

3.  Ribosome profiling = ribosome footprinting = ribo-seq  New technique to measure gene expression  Ribosome density on mRNA transcripts (translation)  Initiating and elongating ribosomes  Genome Wide Information on Protein Synthesis (GWIPS)  Actual protein production in cells  Snapshot of cell  Translational variation under different conditions  Transcription ≠ translation THEORY – Ribosome profiling introduction

4.  Ribosome freezing  Cell lysis  Nuclease digestion  RPF purification  Linker ligation  cDNA  Preparation for PCR  PCR  Sequencing  Analysis Ingolia et al. - Nature protocol (2012) THEORY – Ribosome profiling technique

5.  Initiating ribosomes  Initiation is slow  Initiating codons  protein isoforms  Translation initiation sites (TISs)  Harringtonine, lactimidomycin or pre-treatment with puromycin  Elongating ribosomes  Gene expression  Global and local rates of translation  Identification of novel translated ORFs  Cycloheximide, emetine, chloraminophenicol or thermal freezing (liquid nitrogen) THEORY – Ribosome profiling strategies Ingolia et al. - Cell (2011)

6.  GWIPS-viz is an online tool that provides visualization of ribo-seq data obtained with the ribosome profiling technique  GWIPS-viz is based on UCSC genome browser, developed at the University of California, Santa Cruz  http://gwips.ucc.ie http://gwips.ucc.ie  Project:  Add C. elegans  Add more datasets for other organisms  Add ribosome profiles GWIPS-viz – Introduction

7. GWIPS-viz – Data workflow

8. Profile Translational position within the ribosome A-site: 15nt / P-site: 12nt offset from 5’ RPF Coverage plot RPF GWIPS-viz – Coverage plots vs. profiles

9. GWIPS-viz – Visualizations  Eukaryotes  Prokaryotes  E. coli  B. subtilis OrganismGenome assembly H. sapienshg19 M. musculusmm10 D. reriodanRer7 C. elegansce10 S. CervisiaesacCer3

10. DEMO GWIPS-viz – Visualizations

11. CONCLUSIO N  Addition of C. elegans data  Addition of ribsome coverage plots and mRNA- seq coverage plots  Addition of ribosome profiles that provide more information on translation  Uniform naming convention and color scheme

12. CREDIT S Audrey Michel – supervisor Pavel Baranov – head of LAPTI All members of LAPTI and The Recoding Lab Raphaël Kiekens – promotor