The stem-loop binding protein forms a highly stable and specific complex with the 3 ' stem-loop of histone mRNAs

Replication-dependent histone mRNAs end in a highly conserved 26-nt stem-lo op structure. The stem-loop binding protein (SLBP), an evolutionarily conse rved protein with no known homologs, interacts with the stem-loop in both t he nucleus and cytoplasm and mediates nuclear-cytoplasmic transport as well as 3'-end processing of the pre-mRNA by the U7 snRNP. Here, we examined th e affinity and specificity of the SLBP-RNA interaction. Nitrocellulose filt er-binding experiments showed that the apparent equilibrium dissociation co nstant (K-d) between purified SLBP and the stem-loop RNA is 1.5 nM. Binding studies with a series of stem-loop Variants demonstrated that conserved re sidues in the stem and loop, as well as the 5' and 3' flanking regions, are required for efficient protein recognition. Deletion analysis showed that 3 nt 5' of the stem and 1 nt 3' of the stem contribute to the binding energ y. These data reveal that the high affinity complex between SLBP and the RN A involves sequence-specific contacts to the loop and the top of the stem, as well the base of the stem and its immediate flanking sequences. Together , these results suggest a novel mode of protein-RNA recognition that forms the core of a ribonucleoprotein complex central to the regulation of histon e gene expression.