Development of cap-caged-mRNA for optochemical control of translation in eukaryotic cells

• Applicant: Professorin Dr. Andrea Rentmeister
• Subject Area: Biological and Biomimetic Chemistry ; Biochemistry
• Term: from 2017 to 2022
• Project identifier: Deutsche Forschungsgemeinschaft (DFG) - Project number 390706730

Light is a versatile regulatory element because it is fully orthogonal to most cellular components, noninvasive, controllable in timing and localization to tissues, cells and even subcellular regions. Various optochemical approaches have allowed the control of gene expression at the level of DNA, mRNA, and protein with spatio-temporal control. However, an efficient method to induce translation of exogenous mRNA with spatio-temporal control is missing. We propose an innovative and highly flexible approach for triggering translation of exogenous mRNA with spatio-temporal control. Towards this end, we will develop a chemo-enzymatic route to modify the mRNA cap with photo-cleavable groups and completely block translation of mRNA. We will use the highly promiscuous N7 cap methyltransferase Ecm1 and synthesize novel analogs of the cosubstrate S-adenosyl methionine (AdoMet). Based on our previous work we know that modifications at the N7 position of the cap can completely block translation because they abrogate the delicate interaction between the cap and the translation initiation factor eIF4E. We will then establish light-triggered removal of these caging groups to reconstitute the canonical cap and thus trigger efficient translation of the respective mRNA in mammalian cells.We will first establish our approach in vitro and then implement it in mammalian cells using quantifiable reporter-mRNAs. Finally, we want to find out whether our approach will be suitable to trigger a physiological process (namely apoptosis) with spatio-temporal control, by activating translation of caspase-mRNAs.

• DFG Programme: Research Grants