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The stem-loop binding protein forms a highly stable and specific complex with the 3' stem-loop of histone mRNAs
1Department of Molecular Biophysics and Biochemistry, Yale University, New Haven, Connecticut 06520, USA
RNA (New York, N.Y.)
|February 24, 2001
Summary
Stem-loop binding protein (SLBP) binds histone mRNA stem-loops with high affinity. This interaction is sequence-specific, involving conserved RNA regions and flanking sequences, crucial for regulating histone gene expression.
Area of Science:
- Molecular Biology
- RNA Biology
- Gene Regulation
Background:
- Replication-dependent histone mRNAs feature a conserved 26-nucleotide stem-loop structure.
- Stem-loop binding protein (SLBP) interacts with this structure in both nucleus and cytoplasm.
- SLBP is essential for mRNA transport and 3'-end processing mediated by U7 snRNP.
Purpose of the Study:
- To investigate the affinity and specificity of the interaction between SLBP and its cognate stem-loop RNA.
- To identify the RNA sequence elements critical for SLBP recognition and binding.
Main Methods:
- Nitrocellulose filter-binding assays to determine binding affinity (Kd).
- Binding studies using a series of stem-loop RNA variants.
- Deletion analysis to map critical RNA regions.
Main Results:
- SLBP binds the stem-loop RNA with a high apparent equilibrium dissociation constant (Kd) of 1.5 nM.
- Conserved residues within the stem, loop, and flanking regions are essential for efficient SLBP binding.
- Specific sequence contacts occur at the loop, top and base of the stem, and flanking sequences.
Conclusions:
- SLBP recognizes its target RNA through a complex, sequence-specific mechanism involving multiple RNA elements.
- This high-affinity interaction is central to the formation of a ribonucleoprotein complex.
- The findings elucidate a novel mode of protein-RNA recognition critical for histone gene expression regulation.