Comparative proteomics of the two T. brucei PABPs suggests that PABP2 controls bulk mRNA

Area of Science:

• Molecular Biology
• Cell Biology
• Genetics

Background:

• Poly(A)-binding proteins (PABPs) are crucial regulators of mRNA stability and translation.
• Organisms often possess multiple PABP and translation initiation factor paralogs, suggesting specialized functions.
• Trypanosomes exhibit complex translation machinery with five eIF4G, six eIF4E paralogs, and two PABPs (PABP1, PABP2).

Purpose of the Study:

• To elucidate the distinct protein interaction networks and cellular localization of PABP1 and PABP2 in *Trypanosoma brucei*.
• To understand the differential roles of PABP paralogs in mRNA regulation and stress granule dynamics.

Main Methods:

• Cryo-milling coupled with affinity purification and mass spectrometry was employed to identify protein interactors of PABP1 and PABP2.
• Cellular localization studies, including stress granule and polysome fractionation, were performed under starvation conditions.

Main Results:

• PABP1 exhibited specific interactions with eIF4E4 and eIF4G3, and its associated proteins were largely excluded from starvation stress granules.
• PABP2 displayed promiscuous interactions with numerous translation initiation factors, including eIF4G1, and translocated to stress granules upon starvation.
• PABP1 preferentially associated with small polysomes and a distinct set of RNA-binding proteins, whereas PABP2 interacted with a broader range of proteins and distributed across all polysome fractions.

Conclusions:

• PABP1 likely regulates a small subset of small-sized mRNAs, functioning independently of the major stress granule pathway.
• PABP2 appears to govern bulk mRNA translation, participating in the dynamic relocation to stress granules during starvation.
• The differential interactomes and localization patterns of PABP1 and PABP2 highlight specialized roles in trypanosome mRNA metabolism.