Concerted 2-5A-Mediated mRNA Decay and Transcription Reprogram Protein Synthesis in dsRNA Response

Sneha Rath; Eliza Prangley; Jesse Donovan; Kaitlin Demarest; Yigal Meir; Ned Wingreen; Alexei Korennykh

RNase L cleaves most basal mRNAs with similar rate constants and does not exhibit a preference for longer and AU-rich mRNAs. The uniform mRNA decay is a central feature of 2-5AMD, which is responsible for arrest of all housekeeping proteins rather than just proteins encoded by long and AU-rich mRNAs. We propose that this uniformity indicates that in live cells mRNAs are cleaved in Briggs-Haldane kinetic regime (Methods). Under Briggs-Haldane conditions, RNase L will cleave mRNAs independently of the binding (K m ) and catalytic constants (k cat ) of individual mRNAs, thereby acquiring a mechanism for uniform decay of the transcriptome. A notable feature of Briggs-Haldane regime is that once RNase L encounters an mRNA, it makes a cut before dissociating, i.e. every RNase L-mRNA binding event is productive. In cell extracts, RNase L is sensitive to length and AU content in mRNAs, indicating cleavage under Michaelis-Menten regime. Although it remains to be explained precisely how live . CC-BY 4.0 International license is made available under a The copyright holder for this preprint (which was not peer-reviewed) is the author/funder. It . https://doi.org/10.1101/484675 doi: bioRxiv preprint cells achieve Briggs-Haldane regime, our observations could be explained by ribosomeassisted RNase L access to mRNAs. This model agrees with the recently proposed mechanism for Dom34 rescue of RNase L-ribosome complex, which postulates a translation-dependent mRNA cleavage mechanism (Nogimori et al., 2018) . If ribosome-RNase L recognition (rather than mRNA-RNase L recognition) determines kinetics mRNA decay by RNase L, 2-5AMD would target all actively translating mRNAs. Further, RNase L could dwell on translating ribosomes, which would ensure efficient cleavage of mRNAs and achieve Briggs-Haldane conditions. The decay rate constants are similar, within several-fold, for basal mRNAs and for mRNAs encoding IFNs ( Fig. 4C; 6F ). We show that once this several-fold stability advantage is coupled with the positive feedback of the IFN response, defense mRNAs become desensitized to RNase L. A minimal model with experimentally determined kinetic parameters can account for decay of basal mRNAs and explain how IFNs and ISGs can accumulate to nearly the same levels in WT and RNase L -/cells ( Fig. 7A-B) .