Gene activation guided by nascent RNA-bound transcription factors

Abstract

Technologies for gene activation are valuable tools for the study of gene functions and have a wide range of potential applications in bioengineering and medicine. In contrast to existing methods based on recruiting transcriptional modulators via DNA-binding proteins, we developed a strategy termed Narta (nascent RNA-guided transcriptional activation) to achieve gene activation by recruiting artificial transcription factors (aTFs) to transcription sites through nascent RNAs of the target gene. Using Narta, we demonstrate robust activation of a broad range of exogenous and endogenous genes in various cell types, including zebrafish embryos, mouse and human cells. Importantly, the activation is reversible, tunable and specific. Moreover, Narta provides better activation potency of some expressed genes than CRISPRa and, when used in combination with CRISPRa, has an enhancing effect on gene activation. Quantitative imaging illustrated that nascent RNA-directed aTFs could induce the high-density assembly of coactivators at transcription sites, which may explain the larger transcriptional burst size induced by Narta. Overall, our work expands the gene activation toolbox for biomedical research.

Stochastic Modeling of Biophysical Responses to Perturbation

“…In each activation condition, we found that burst sizes of the corresponding target genes were increased (with, on average, log 2 FCs greater than 2) (Fig. 2e left), mirroring burst size observations discussed in the Narta activation protocol, which recruited artificial transcription factors to the transcription site [40]. In comparison, average splicing and degradation rate FCs were near zero (Fig.…”

Exploring tools for studying asymmetric cell division in the zebrafish developing telencephalon

“…Therefore, the low efficiency of the construct expression with intronic MS2 repeats cannot be explained by changes in mRNA splicing dynamics. One study conducted in zebrafish used the MS2-MCP system as a way of recruiting artificial transcription factors to nascent mRNAs and reported that constructs with an intronic MS2 position function more efficiently than 3' ones (Liang et al, 2022). Nevertheless, in our experiments, intronic MS2 positioning did not yield the expected results, potentially due to other interfering factors discussed here.…”