DZNep-induced single point mutation (M236I) in poxviral 2'-O-methyltransferase enhances mRNA stability and translation efficiency

VERMA, A.; Jangir, R. K.; Kamboj, H.; Kumar, G.; Meena, V. K.; Sharma, V.; Bhattacharaya, T. K.; Tripathi, B. N.; Sharma, S.; Kumar, N.

doi:10.1101/2025.03.10.642379

bioRxiv · 10.1101/2025.03.10.642379

DZNep-induced single point mutation (M236I) in poxviral 2'-O-methyltransferase enhances mRNA stability and translation efficiency

VERMA, A.·Jangir, R. K.·Kamboj, H.·Kumar, G.·Meena, V. K.·Sharma, V.·Bhattacharaya, T. K.·Tripathi, B. N.·Sharma, S.·Kumar, N.

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Abstract

Poxviruses encode two highly conserved S-adenosylmethionine (SAM)-dependent methyltransferases for mRNA capping. While D1L facilitates translation via cap-0 formation, the role of VP39-mediated cap-1 modification has been canonically restricted to shield viral mRNA from cytoplasmic pattern-recognition receptors (PRRs). Here, we uncover a novel function of VP39 by demonstrating that prolonged exposure to the SAM cycle inhibitor DZNep selected a virus escape mutant harbouring a point mutation (M236I) in VP39. VP39_mut bypasses S-adenosylhomocysteine (SAH)-mediated feedback inhibition, enhancing 2-O-methylation of viral transcripts and promoting their association with active translation machinery. Consequently, VP39_mut significantly improves viral protein translation, replicative fitness, and yields across species. Notably, VP39_mut-capped in vitro transcribed (IVT) mRNA exhibited [~]2.7-fold higher protein expression compared to VP39_wt, the standard tool for IVT applications. These findings reveal a unique resistance mechanism and provide critical insights into poxvirus mRNA biology while underscoring the potential of optimized VP39 for advancing mRNA-based therapeutics and vaccine production.

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VERMA, A., Jangir, R. K., Kamboj, H., Kumar, G., Meena, V. K., Sharma, V., Bhattacharaya, T. K., Tripathi, B. N., Sharma, S., Kumar, N.. 2025-03-11. DZNep-induced single point mutation (M236I) in poxviral 2'-O-methyltransferase enhances mRNA stability and translation efficiency. https://doi.org/10.1101/2025.03.10.642379

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