Understanding How Two Similar RNA Binding Domains of Paralogous Proteins Mediate Different Protein‐Protein Interactions

Abstract

Polypyrimidine Tract Binding Protein (PTBP) is an RNA binding protein that has a high affinity for pyrimidine rich regions of RNA sequences. Studies have shown that it acts primarily as a negative regulator of alternative pre‐mRNA splicing events (ASEs); however, it has been shown in some cases to enhance ASEs. Improper regulation of ASEs is linked to neurodegenerative diseases and cancer. Two well studied paralogs of PTBP are PTBP1 and PTBP2. The two proteins demonstrate different levels of splicing repression activity despite sharing high amino acid sequence identity (74%) and similar domain arrangement consisting of four RNA recognition motifs (RRMs) connected by three linker sequences and an N‐terminal region. Recent studies generating chimeras of the two proteins demonstrated that multiple regions of PTBP1 can confer splicing repression activity when integrated in the PTBP2 backbone. These studies also highlighted that despite the high amino acid sequence identity, the RRM 2 region of the two proteins demonstrate different affinities for a partner protein, Raver 1. This protein is required for complete PTB‐mediated splicing repression of the α‐tropomyosin exon 3 sequence. The PTBP‐Raver1 interaction motif has been identified, and PTBP1 and PTBP2 share 87% amino acid sequence identity within their respective Raver1 interaction motifs. These results suggest that additional factors such as post‐translational modifications (PTMs) might influence the PTBP‐Raver1 interaction and in turn, splicing repression activity. To test this, we assayed PTBP1 RRM2 for PTMs via immunoprecipitation followed by mass spectrometry and have identified several ubiquitin modifications. Next, we plan to examine whether the ubiquitin modifications in PTBP1 RRM2 contributes to the Raver1 interaction specifically, as well as to other protein‐protein interactions required for splicing repression activity.Support or Funding InformationThis work was supported by a Maximizing Access to Research Careers grant to CSUF from the National Institutes of Health [5T34GM008612‐21]This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.