Stem-loop binding protein facilitates 3 '-end formation by stabilizing U7 snRNP binding to histone pre-mRNA

The 3' end of histone mRNA is formed by an endonucleolytic cleavage of the primary transcript after a conserved stem-loop sequence. The cleavage react ion requires at least two trans-acting factors: the stem-loop binding prote in (SLBP), which binds the stem-loop sequence, and the U7 snRNP that intera cts with a sequence downstream from the cleavage site. Removal of SLBP from a nuclear extract abolishes 3'-end processing, and the addition of recombi nant SLBP restores processing activity of the depleted extract. To determin e the regions of human SLBP necessary for 3' processing, various deletion m utants of the protein were tested for their ability to complement the SLBP- depleted extract. The entire N-terminal domain and the majority of the C-te rminal domain of human SLBP are dispensable for processing. The minimal pro tein that efficiently supports cleavage of histone pre-mRNA consists of 93 amino acids containing the 73-amino-acid RNA-binding domain and 20 amino ac ids located immediately next to its C terminus. Replacement of these 20 res idues with an unrelated sequence in the context of the full-length SLBP red uces processing > 90%. Coimmunoprecipitation experiments with the anti-SLBP antibody demonstrated that SLBP and U7 snRNP form a stable complex only in the presence of pre-mRNA substrates containing a properly positioned U7 sn RNP binding site. One role of SLBP is to stabilize the interaction of the h istone pre-mRNA with U7 snRNP.