Structural basis of pre-mRNA recognition by the human cleavage factor Im complex

Area of Science:

• Molecular Biology
• Structural Biology
• Biochemistry

Background:

• Cleavage factor I(m) (CF I(m)) is crucial for pre-mRNA 3'-end processing.
• CF I(m) recognizes specific sequences upstream of poly(A) sites.
• The complex comprises a 25 kDa subunit (CF I(m)25) and larger subunits (CF I(m)59, CF I(m)68, CF I(m)72).

Purpose of the Study:

• To determine the crystal structure of human CF I(m) complexed with RNA.
• To elucidate the structural basis of CF I(m)-RNA recognition and binding.
• To understand the role of CF I(m)68 in pre-mRNA binding.

Main Methods:

• X-ray crystallography of human CF I(m)25 and CF I(m)68RRM.
• Crystallography of the CF I(m)-RNA complex.
• Point mutation and kinetic analyses.

Main Results:

• The crystal structure reveals a heterotetrameric complex of CF I(m)25 dimer and two CF I(m)68RRM molecules.
• Two UGUAA RNA sequences bind in an anti-parallel orientation to the heterotetramer.
• CF I(m)68RRM binds flanking sequences and enhances overall RNA-binding affinity.

Conclusions:

• The study provides a structural mechanism for CF I(m) binding two UGUAA elements in pre-mRNA.
• CF I(m)68 plays a critical role in the sequence-dependent binding of CF I(m) to pre-mRNA.
• These findings offer insights into the regulation of pre-mRNA polyadenylation.