Light-control of cap methylation and mRNA translation via genetic code expansion of Ecm1. Issue 12 (12th February 2021)

• Record Type: Journal Article
• Authors: Reichert, Dennis; Mootz, Henning D.; Rentmeister, Andrea
• Is Part Of: Chemical science. Volume 12:Issue 12(2021)
• Page Start: 4383
• Page End: 4388
• Publication Date: 2021-02-12
• Subjects: Chemistry -- Periodicals; 540.5
• DOI: 10.1039/d1sc00159k
• Languages: English
• ISSNs: 2041-6520

Abstract

Abstract : A tyrosine in the active site of the 5′ cap methyltransferase Ecm1 was photocaged. Translation of mRNA could be triggered by light in eukaryotic cell lysate. 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.