Dissecting the RNA-binding capacity of the multi-RRM protein Rrm4 essential for endosomal mRNA transport

Area of Science:

• Molecular Biology
• Genetics
• Cell Biology

Background:

• RNA-binding proteins (RBPs) are crucial for gene regulation, interacting with messenger RNA (mRNA) networks via multiple RNA-binding domains (RBDs).
• Understanding how individual RBDs contribute to RBP function and transcriptome-wide interactions is essential but remains challenging.

Purpose of the Study:

• To investigate the specific roles of the three RNA recognition motifs (RRMs) within the endosomal mRNA transporter Rrm4 in Ustilago maydis.
• To elucidate how Rrm4's modular RBDs shape transcriptome-wide RNA binding and influence mRNA fate during polar growth.

Main Methods:

• Comparative mutant-based individual-nucleotide resolution crosslinking and immunoprecipitation (iCLIP2) was employed to map RRM-specific binding sites.
• Integration of transcriptome-wide RNA binding data with transcriptomics (RNA-seq) was performed.

Main Results:

• An extensive inventory of RRM-specific binding sites for Rrm4 was identified.
• While the third RRM domain governs most binding sites, RRM1 and RRM2 are critical for functionally essential interactions, involving complex RBD interplay.
• Rrm4 binding regulates mRNA abundance, linking endosomal transport to mRNA stability and defining RNA regulons impacting mitochondrial activity, polarity, and cell wall remodeling.

Conclusions:

• The modular RNA binding of Rrm4, through differential use of its RRMs, distinguishes functional from accessory binding sites.
• This mechanism dictates mRNA fate and is critical for cellular processes like polar growth in Ustilago maydis.