A systematic survey of PRMT interactomes reveals the key roles of arginine methylation in the global control of RNA splicing and translation.

Thousands of proteins undergo arginine methylation, a widespread post-translational modification catalyzed by several protein arginine methyltransferases (PRMTs). However, global understanding of their biological functions is limited due to the lack of a complete picture of the catalytic network for each PRMT. Here, we systematically identified interacting proteins for all human PRMTs and demonstrated their functional importance in mRNA splicing and translation. We demonstrated significant overlapping of interactomes of human PRMTs with the known methylarginine-containing proteins. Different PRMTs are functionally redundant with a high degree of overlap in their substrates and high similarities between their putative methylation motifs. Importantly, RNA-binding proteins involved in regulating RNA splicing and translation contain highly enriched arginine methylation regions. Moreover, inhibition of PRMTs globally alternates alternative splicing (AS) and suppresses translation. In particular, ribosomal proteins are extensively modified with methylarginine, and mutations in their methylation sites suppress ribosome assembly, translation, and eventually cell growth. Collectively, our study provides a global view of different PRMT networks and uncovers critical functions of arginine methylation in regulating mRNA splicing and translation.

Expanding RNA binding specificity and affinity of engineered PUF domains.

Specific manipulation of RNA is necessary for the research in biotechnology and medicine. The RNA-binding domains of Pumilio/fem-3 mRNA binding factors (PUF domains) are programmable RNA binding scaffolds used to engineer artificial proteins that specifically modulate RNAs. However, the native PUF domains generally recognize 8-nt RNAs, limiting their applications. Here, we modify the PUF domain of human Pumilio1 to engineer PUFs that recognize RNA targets of different length. The engineered PUFs bind to their RNA targets specifically and PUFs with more repeats have higher binding affinity than the canonical eight-repeat domains; however, the binding affinity reaches the peak at those with 9 and 10 repeats. Structural analysis on PUF with nine repeats reveals a higher degree of curvature, and the RNA binding unexpectedly and dramatically opens the curved structure. Investigation of the residues positioned in between two RNA bases demonstrates that tyrosine and arginine have favored stacking interactions. Further tests on the availability of the engineered PUFs in vitro and in splicing function assays indicate that our engineered PUFs bind RNA targets with high affinity in a programmable way.