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Isolation of Cognate RNA-protein Complexes from Cells Using Oligonucleotide-directed Elution
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Isolation of Cognate RNA-protein Complexes from Cells Using Oligonucleotide-directed Elution

Published on: January 16, 2017

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Upstream of N-Ras C-terminal cold shock domains mediate poly(A) specificity in a novel RNA recognition mode and bind

Nele Merret Hollmann1,2, Pravin Kumar Ankush Jagtap1,3, Johanna-Barbara Linse4,5

  • 1Structural and Computational Biology Unit, EMBL Heidelberg, Meyerhofstraße 1, 69117 Heidelberg, Germany.

Nucleic Acids Research
|January 23, 2023
PubMed

Insights

This study reveals how the Upstream of N-Ras (Unr) protein uses multiple RNA binding domains to achieve specific RNA targeting. Structural insights show complex interactions beyond classical models, guiding future research on protein partnerships in cellular functions.

Area of Science:

  • Molecular Biology
  • Structural Biology
  • Biochemistry

Background:

  • RNA binding proteins (RBPs) utilize multiple RNA binding domains (RBDs) for enhanced RNA target specificity and affinity.
  • The intricate mechanisms governing multi-RBD target recognition are not fully understood.

Purpose of the Study:

  • To elucidate how multiple RBDs orchestrate target specificity using the multidomain RBP, Upstream of N-Ras (Unr).
  • To provide high-resolution structural insights into Unr-RNA and Unr-protein interactions.

Main Methods:

  • X-ray crystallography to determine the structure of Unr's cold-shock domains (CSDs) bound to poly(A) RNA.
  • Structural studies to identify interaction surfaces between Unr's N-terminal and C-terminal regions with poly(A)-binding protein (pAbp).
  • Mutational analyses to validate observed interactions.

Main Results:

  • A crystal structure revealed Unr's CSDs binding poly(A) RNA through mechanisms extending beyond canonical ππ-stacking.
  • Identified multiple interaction surfaces between Unr and pAbp, highlighting a complex protein-protein interaction network.
  • Mutational data confirmed the functional relevance of the elucidated interaction sites.

Conclusions:

  • Unr employs a sophisticated multi-RBD strategy for RNA recognition, involving novel binding modes.
  • The structural and mutational data provide a foundation for understanding the collaborative roles of Unr and pAbp in cellular processes.
  • These findings offer insights into the broader principles of multi-domain RBP function and RNA regulation.

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