Abstract
In metazoans, replication-dependent histone mRNAs, unlike all other mRNAs, are not polyadenylated but instead terminate with a unique, highly conserved sequence containing a 6-bp stem and a 4-base loop (1). This relatively short, 26-nucleotide sequence has multiple functions in metabolism of all replication-dependent histone mRNAs, including their nuclear export (2,3) and localization to polyribosomes (4,5), as well as regulation of their half-life in the cytoplasm (6). In metabolism of all other mRNAs, these functions are mediated by the poly(A) tail. Additionally, the histone stem-loop sequence is required for efficient 3′ end cleavage of replication-dependent histone pre-mRNAs, leading to the formation of mature histone mRNAs (7,8). Critical features of the 16-nucleotide stem-loop structure and the 5-nucleotide flanking sequences are shown in Fig. 1 A. The two GC base pairs at the base of the stem and the UA base pair at the top of the stem are invariant. Moreover, there are virtually always uridines in the first and third nucleotides of the loop. The only known exception is the loop of Caenorhabditis elegans histone mRNA, which in the first position contains cytidine rather than uridine (9). Both flanking sequences contain mostly adenines and cytidines, with CCAAA consensus sequence on the 5′ side and ACCA or ACCCA the consensus on the 3′ side.
The last 26 nucleotides of replication-dependent histone mRNAs are recognized by the SLBP. (A) Consensus sequence for the 16-nucleotide stem-loop structure and the 5-nucleotide flanking regions at the 3′ end of histone mRNAs. The invariant nucleotides are boxed. (B) The 3′ end of the mouse H2a-614 histone mRNA and the reverse-stem mutant (RS) unable to bind SLBP. Altered nucleotides are boxed. (C) Binding of SLBP from the nuclear extract of the mouse myeloma cells to the radiolabeled RNA containing the H2a -614 wild-type stem-loop sequence, as detected by mobility shift assay. A slowly migrating complex of SLBP and the RNA probe (lane 2) can be competed by 50-fold excess of the cold wild-type RNA (lane 3) but not by a 1000-fold excess of the RS mutant, which is unable to bind SLBP (lane 4). The probe is shown in lane 1.
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Dominski, Z., Marzluff, W.F. (2001). Three-Hybrid Screens for RNA-Binding Proteins. In: MacDonald, P.N. (eds) Two-Hybrid Systems. Methods in Molecular Biology, vol 177. Humana Press. https://doi.org/10.1385/1-59259-210-4:291
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DOI: https://doi.org/10.1385/1-59259-210-4:291
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