Replication-dependent histone mRNAs end in a highly
conserved 26-nt stem-loop structure. The stem-loop binding
protein (SLBP), an evolutionarily conserved protein with
no known homologs, interacts with the stem-loop in both
the 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 the affinity and specificity
of the SLBP–RNA interaction. Nitrocellulose filter-binding
experiments showed that the apparent equilibrium dissociation
constant (Kd) between purified SLBP
and the stem-loop RNA is 1.5 nM. Binding studies with a
series of stem-loop variants demonstrated that conserved
residues 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 energy. These data reveal that the high affinity
complex between SLBP and the RNA 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 histone gene expression.