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Structural landscape of Microprocessor-mediated processing of pri-let-7 miRNAs

Ankur Garg
Ankur Garg

Author interview: Ankur Garg, Dr.rer.nat. is a postdoctoral scientist working with Howard Hughes Medical Institute (HHMI) at Cold Spring Harbor Laboratory (CSHL) in New York, USA. He studies the molecular machines involved in miRNA biogenesis and RNA interference (RNAi). Ankur uses Cryo-EM and X-ray crystallography to determine protein and protein-RNA complex structures, and complement structural findings with different biochemical & biophysical methods.

Lab: Leemor Joshua-Tor, Ph.D., Cold Spring Harbor Laboratory, New York, USA.

MicroRNA (miRNA) biogenesis is initiated upon cleavage of a primary miRNA (pri-miRNA) hairpin by the Microprocessor (MP), composed of the Drosha RNase III enzyme and its partner DGCR8. Pri-miRNAs have multiple sequence motifs that affect MP recognition, fidelity, and efficiency.

What was the core problem you aimed to solve with this research?

MicroRNA (miRNA) biogenesis is initiated upon cleavage of a primary miRNA (pri-miRNA) hairpin by the Microprocessor (MP), composed of the Drosha RNase III enzyme and its partner DGCR8. Pri-miRNAs have multiple sequence motifs that affect MP recognition, fidelity, and efficiency. We aimed to understand how human MP utilizes these motifs to specifically recognize different pri-miRNAs substrates.

Cartoon and space-fill representation of cryo-EM structures of Microprocessor in complex with different pri-let-7 miRNA substrates.
Cartoon and space-fill representation of cryo-EM structures of Microprocessor in complex with different pri-let-7 miRNA substrates.

How did you go about solving this problem?

We performed cryo-electron microscopy (cryo-EM) and biochemical studies of several let-7 family pri-miRNAs in complex with human MP. We found that MP has the structural plasticity to accommodate a range of pri-miRNAs. These structures revealed key features of the 5′ UG sequence motif, more comprehensively represented as the “flipped U with paired N” (fUN) motif. Our analysis explains how cleavage of class-II pri-let-7 members harboring a bulged nucleotide generates a non-canonical precursor with a 1-nt 3′ overhang. Finally, the MP-SRSF3-pri-let-7f1 structure reveals how SRSF3 contributes to MP fidelity by interacting with the CNNC motif and Drosha’s Piwi/Argonaute/Zwille (PAZ)-like domain. Overall, our study shed light on the mechanisms for flexible recognition, accurate cleavage, and regulated processing of different pri-miRNAs by MP.

Series of cryo-EM structures of the Microprocessor with different pri-let-7 miRNA substrates allowed us to comprehensively define sequence features conserved in pri-miRNAs. — Leemor Joshua-Tor, Ph.D.

What was the exciting moment during your research?

The most exciting moment in cryo-EM research is to see the protein/protein-RNA complex particles on the micrograph, and my eyes always shine bright when those micrographs eventually generate a 3D cryoEM map. Specifically, in this work when I saw the 3D map density for the pri-miRNA helix for the first time, was the most exciting moment.

Research article: The structural landscape of Microprocessor-mediated processing of pri-let-7 miRNAs. Molecular Cell, Volume 84, Issue 21, 4175 – 4190.e6. https://www.cell.com/molecular-cell/fulltext/S1097-2765(24)00741-X.

Read more author interviews here: https://biopatrika.com/category/academia/researcher-spotlight/

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