Structural and functional insights into the fly microRNA biogenesis factor Loquacious

Area of Science:

• Molecular Biology
• Structural Biology
• Genetics

Background:

• MicroRNA (miRNA) biogenesis relies on Dicer enzymes for precursor processing.
• Double-stranded RNA-binding proteins (dsRBPs) are essential cofactors for Dicer activity.
• In Drosophila, the dsRBP Loquacious (Loqs) partners with Dicer1 (dmDcr1) to mediate miRNA production.

Purpose of the Study:

• To elucidate the structural basis of the interaction between Loqs and dmDcr1.
• To identify the specific domains and elements within Loqs responsible for dmDcr1 binding.
• To investigate the potential role of Loqs dimerization in miRNA processing.

Main Methods:

• X-ray crystallography was employed to determine the structure of the third dsRBD of Loqs.
• Biochemical assays were used to map the interaction interfaces between Loqs and dmDcr1.
• Site-directed mutagenesis was performed to probe the function of the Loqs dimerization interface.

Main Results:

• The crystal structure of the Loqs dsRBD3 revealed specific interaction surfaces for dmDcr1.
• A linker region N-terminal to dsRBD3 significantly contributes to dmDcr1 binding affinity.
• Loqs dsRBD3 forms homodimers, and mutations at the interface disrupt dmDcr1 interaction.
• Loqs binds dmDcr1 as a monomer, utilizing the dimerization surface, suggesting alternative dimerization states.

Conclusions:

• The structural and functional data define key elements in Loqs for dmDcr1 interaction and miRNA processing.
• Loqs dimerization is a critical feature, potentially regulating its interaction with dmDcr1.
• Conserved structural motifs suggest that dsRBP dimerization may be a conserved mechanism in gene silencing pathways.