cb0c00205_si_001.pdf (4.7 MB)
Multiplexed Photoactivation of mRNA with Single-Cell Resolution
journal contribution
posted on 2020-06-12, 19:35 authored by Dongyang Zhang, Shuaijiang Jin, Xijun Piao, Neal K. DevarajWe
demonstrate sequential optical activation of two types of mRNAs
in the same mammalian cell through the sequential photocleavage of
small molecule caging groups (“photocages”) tethered
to the 5′-untranslated region (5′-UTR) of mRNAs. Synthetic
photocages were conjugated onto target mRNA using RNA-TAG, an enzymatic
site-specific RNA modification technique. Translation of mRNA was
severely reduced upon conjugation of the photocages onto the 5′-UTR.
However, subsequent photorelease of the cages from the mRNA transcript
triggered activation of translation with single-cell spatiotemporal
resolution. To achieve sequential photoactivation of two mRNAs in
the same cell, we synthesized a pair of photocages that can be selectively
cleaved from mRNA upon photoirradiation with different wavelengths
of light. Sequential photoactivation of two mRNAs enabled precise
optical control of translation of two unique transcripts. We believe
that this modular approach to precisely and rapidly control gene expression
will serve as a powerful tool in future biological studies that require
controlling translation of multiple transcripts with high spatiotemporal
resolution.