Effects of an RNA chaperone on mutation tolerance

Due to their intrinsic thermodynamic properties, RNA can misfold easily in cells. One way to mitigate RNA misfolding is through the actions of RNA chaperones, which bind and unwind structured RNA molecules and thereby offer opportunities for these misfolded species to refold properly. Such rescue activity has implications for the fitness effects of individual mutations-- at least mutations that compromise RNA folding or structure might be buffered by RNA chaperones. However, little is known about the rules governing such mutation buffering. Here, we describe how a model RNA chaperone, the DEAD-box RNA helicase CYT-19, affects the fitness effects of mutations in a model structured RNA, the Tetrahymenagroup I intron, whose self-splicing activity is dependent on its structure. We performed deep mutational scanning on the P1ex region of the intron which is critical for its self-splicing activity, and assayed differential splicing activity of all possible P1ex mutants in the presence and absence of CYT-19 to identify mutations that are buffered by RNA chaperone activity. I will discuss the properties of the chaperone-dependent and chaperone-independent mutation pools. Our results highlight that, to understand RNA robustness in vivo, we need to consider how mutational fitness effects are modulated by RNA chaperones and other trans-acting factors.