Lars Scharff - Regulation and fine-tuning of protein synthesis in chloroplasts

Protein synthesis in chloroplasts contributes up to 50% of the total plant proteome, and is a major regulatory level of gene expression in chloroplasts. The protein synthesis machinery in plastids consists of an interesting mixture of strongly conserved bacterial features and rapidly evolving eukaryotic proteins. I am interested in unravelling the mechanisms regulating translation, for which we use different omics technologies.

It is well known that translation initiation regulation is executed by RNA binding proteins, but much less is known is about the mechanisms, by which the binding of these proteins influences translation. To address this question, we analyse to which extend regulation of translation initiation is executed by altering mRNA secondary structure. Selective 2′ Hydroxyl Acylation analysed by Primer Extension (SHAPE) was used to probe in vivo RNA structure.

Furthermore we study what determines the speed of translation elongation, and which processes are influenced by ribosome pausing. This study is based on ribosome profiling data, i.e. sequencing of ribosome footprints. Our results show that mRNA secondary structure, Shine-Dalgarno sequences in coding regions, and positively charged amino acids in the nascent peptide chains work together to cause ribosome pausing. Ribosome pausing likely influences transmembrane protein folding and co-factor incorporation. The features influencing pausing are conserved in land plants, but less so in parasitic plants indicating co-evolution of the regulation of the speed of translation elongation and assembly of photosynthetic complexes.