CryoEM of RUVBL1-RUVBL2-ZNHIT2, a complex that interacts with pre-mRNA-processing-splicing factor 8

Area of Science:

• Molecular Biology
• Protein Biochemistry
• Structural Biology

Background:

• Biogenesis of the U5 small nuclear ribonucleoprotein (snRNP) is critical for cellular function.
• PRPF8, a key U5 snRNP component, requires the HSP90 chaperone and the R2TP cochaperone complex (containing RUVBL1 and RUVBL2 ATPases) for its assembly.
• Additional factors regulating R2TP during PRPF8 biogenesis remain incompletely understood.

Purpose of the Study:

• To investigate the role of ZNHIT2 in regulating the R2TP chaperone during PRPF8 biogenesis.
• To elucidate the structural and functional impact of ZNHIT2 on the R2TP complex.

Main Methods:

• Biochemical assays
• Interaction mapping
• Mass spectrometry
• Cryo-electron microscopy (cryo-EM)

Main Results:

• ZNHIT2 forms a stable complex with the R2TP cochaperone via direct interaction with RUVBL1-RUVBL2 ATPases.
• Cryo-EM analysis revealed that ZNHIT2 binding alters the conformation and nucleotide state of RUVBL1-RUVBL2, modulating its ATPase activity.
• PRPF8 directly interacts with R2TP, and this complex can incorporate ZNHIT2, ECD, and AAR2, highlighting a network of assembly factors.

Conclusions:

• ZNHIT2 is a novel regulator of the R2TP chaperone complex during U5 snRNP biogenesis.
• ZNHIT2 influences PRPF8 assembly by modulating R2TP structure and activity.
• These findings reveal ZNHIT2's integral role within a network of factors coordinating PRPF8 biogenesis and R2TP-HSP90 chaperone function.