Post-Translational Modifications in Polypyrimidine Tract Binding Proteins PTBP1 and PTBP2

Area of Science:

• Molecular Biology
• Gene Regulation
• Post-Translational Modifications

Background:

• RNA binding proteins are crucial for regulating gene expression through alternative pre-mRNA splicing.
• Members of RNA binding protein families often share structural similarities but exhibit distinct tissue-specific functions and target different exons.
• The mechanisms by which highly similar paralogs achieve differential splicing outcomes remain largely unknown.

Purpose of the Study:

• To investigate post-translational modifications (PTMs) in paralogous RNA binding proteins PTBP1 and PTBP2 using mass spectrometry.
• To identify specific phosphorylation and acetylation sites that may dictate the differential splicing activities of PTBP1 and PTBP2.

Main Methods:

• Mass spectrometry was employed to analyze post-translational phosphorylation and acetylation modifications.
• Analysis focused on PTBP1 and PTBP2 within splicing reaction mixtures to capture relevant modifications.
• Identification of distinct and overlapping modification sites across protein regions, including RNA recognition motifs (RRMs).

Main Results:

• PTBP1 and PTBP2 exhibit numerous distinct phosphorylation sites, primarily in their N-terminal, linker 1, and linker 2 regions.
• Acetylation patterns show significant overlap in the RRMs, with unique acetylation sites identified in PTBP1 RRM2 and RRM3.
• Acetylation of lysine residues within the nuclear localization sequence of PTBP2 was observed.

Conclusions:

• Significant differences in post-translational modification profiles exist between PTBP1 and PTBP2 paralogs.
• These distinct PTMs, particularly phosphorylation and acetylation, are likely key determinants of their differential splicing activities.
• The findings provide insights into how protein paralogs achieve functional specificity in gene regulation.