Light-control of cap methylation and mRNA translation via genetic code expansion of Ecm1
Open Access
- 8 February 2021
- journal article
- research article
- Published by Royal Society of Chemistry (RSC) in Chemical Science
- Vol. 12 (12), 4383-4388
- https://doi.org/10.1039/d1sc00159k
Abstract
Gene expression is tightly regulated in all domains of life, with post-transcriptional regulation being more pronounced in higher eukaryotes. Optochemical and optogenetic approaches enable the actuation of many underlying processes by light, which is an excellent tool to exert spatio-temporal control. However, light-mediated control of eukaryotic mRNA processing and the respective enzymes has not been reported. We used genetic code expansion to install a photo-caged tyrosine (Y) in the active site of the cap methyltransferase Ecm1. This enzyme is responsible for guanine N7 methylation of the 5′ cap, which is required for translation. Substituting Y284 with the photocaged ortho-nitrobenzyl-tyrosine (ONBY) almost completely abrogated the methylation activity of Ecm1. Irradiation with light removed the ONB group, restoring the native tyrosine and Ecm1 activity, yielding up to 97% conversion of the minimal substrate GpppA within 60 min after activation. Using luciferase- and eGFP-mRNAs as reporters, we could show that light actuates translation by inducing activation of Ecm1 ONBY284 in a eukaryotic in vitro translation system.Funding Information
- Deutsche Forschungsgemeinschaft (RE2796/6-1, RE2796/7-1, SFB858, SFB1459)
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