Reading ADP-ribosylation signaling using chemical biology and interaction proteomics.

ADP-ribose (ADPr) readers are essential components of ADP-ribosylation signaling, which regulates genome maintenance and immunity. The identification and discrimination between monoADPr (MAR) and polyADPr (PAR) readers is difficult because of a lack of suitable affinity-enrichment reagents. We synthesized well-defined ADPr probes and used these for affinity purifications combined with relative and absolute quantitative mass spectrometry to generate proteome-wide MAR and PAR interactomes, including determination of apparent binding affinities. Among the main findings, MAR and PAR readers regulate various common and distinct processes, such as the DNA-damage response, cellular metabolism, RNA trafficking, and transcription. We monitored the dynamics of PAR interactions upon induction of oxidative DNA damage and uncovered the mechanistic connections between ubiquitin signaling and ADP-ribosylation. Taken together, chemical biology enables exploration of MAR and PAR readers using interaction proteomics. Furthermore, the generated MAR and PAR interaction maps significantly expand our current understanding of ADPr signaling.

Decoupled degradation and translation enables noise modulation by poly(A) tails.

Poly(A) tails are crucial for mRNA translation and degradation, but the exact relationship between tail length and mRNA kinetics remains unclear. Here, we employ a small library of identical mRNAs that differ only in their poly(A)-tail length to examine their behavior in human embryonic kidney cells. We find that tail length strongly correlates with mRNA degradation rates but is decoupled from translation. Interestingly, an optimal tail length of ∼100 nt displays the highest translation rate, which is identical to the average endogenous tail length measured by nanopore sequencing. Furthermore, poly(A)-tail length variability-a feature of endogenous mRNAs-impacts translation efficiency but not mRNA degradation rates. Stochastic modeling combined with single-cell tracking reveals that poly(A) tails provide cells with an independent handle to tune gene expression fluctuations by decoupling mRNA degradation and translation. Together, this work contributes to the basic understanding of gene expression regulation and has potential applications in nucleic acid therapeutics.