Delineating the kinetic RNA interactome of nuclear exosome adaptor complexes

The human genome is ubiquitously transcribed into far more RNA than is immediately needed. Consequently, RNA turnover becomes critical for keeping our cells healthy.

The nuclear RNA exosome degrades the majority of short-lived RNA species within cell nuclei and is hereby the gatekeeper of an enormous RNA synthesis output.

To perform this essential task, the exosome employs so-called ‘adaptor complexes’ (ExoACs), which contribute to target specificity.

While the trimeric nuclear exosome targeting (NEXT) complex is specifically required for the degradation of short RNA transcripts, the poly(A) tail exosome targeting (PAXT) connection recruits the exosome to longer polyadenylated RNAs. Interestingly, and in contrast to NEXT, PAXT exhibits a rather complex nature.

Besides the integral component RNA helicase MTR4 and the large scaffolding protein ZFC3H1, the proteins PABPN1, ZC3H3, and RBM26/27 participate in the RNA-targeting of PAXT.

How assembly of all these proteins is achieved to commit a polyadenylated RNA for decay and how it is balanced with other nuclear RNA metabolic processes to avoid untimely decay of mRNA is unknown.Here, I will track the interaction of the RNA-binding components of NEXT and PAXT with newly synthesized RNA in human cells using a novel and cutting-edge temporally resolved CLIP (T-CLIP) methodology, which combines nascent RNA labeling using photoactivatable ribonucleoside analogue 4-thiouridine with the time course of UV crosslinking of protein-RNA complexes.

Since the T-CLIP approach allows us to link ExoAC interaction profiles to RNA synthesis and turnover data, I will define the positioning and timing of ExoAC component interactions with nuclear RNA in real-time to understand how nuclear exosome identify and eliminate nonfunctional RNA.

In achieving my aim, I will critically further our understanding of how the cells in our bodies manage to quality control their genetic information through the post-transcriptional control of RNA levels.