Replication stress-induced alternative mRNA splicing alters properties of the histone RNA-binding protein HBP/SLBP: a key factor in the control of histone gene expression

Abstract

Animal replication-dependent histone genes produce histone proteins for the packaging of newly replicated genomic DNA. The expression of these histone genes occurs during S phase and is linked to DNA replication via S-phase checkpoints. The histone RNA-binding protein HBP/SLBP (hairpin-binding protein/stem-loop binding protein), an essential regulator of histone gene expression, binds to the conserved hairpin structure located in the 3′UTR (untranslated region) of histone mRNA and participates in histone pre-mRNA processing, translation and histone mRNA degradation. Here, we report the accumulation of alternatively spliced HBP/SLBP transcripts lacking exons 2 and/or 3 in HeLa cells exposed to replication stress. We also detected a shorter HBP/SLBP protein isoform under these conditions that can be accounted for by alternative splicing of HBP/SLBP mRNA. HBP/SLBP mRNA alternative splicing returned to low levels again upon removal of replication stress and was abrogated by caffeine, suggesting the involvement of checkpoint kinases. Analysis of HBP/SLBP cellular localization using GFP (green fluorescent protein) fusion proteins revealed that HBP/SLBP protein and isoforms lacking the domains encoded by exon 2 and exons 2 and 3 were found in the nucleus and cytoplasm, whereas HBP/SLBP lacking the domain encoded by exon 3 was predominantly localised to the nucleus. This isoform lacks the conserved region important for protein–protein interaction with the CTIF [CBP80/20 (cap-binding protein 80/20)]-dependent initiation translation factor and the eIF4E (eukaryotic initiation factor 4E)-dependent translation factor SLIP1/MIF4GD (SLBP-interacting protein 1/MIF4G domain). Consistent with this, we have previously demonstrated that this region is required for the function of HBP/SLBP in cap-dependent translation. In conclusion, alternative splicing allows the synthesis of HBP/SLBP isoforms with different properties that may be important for regulating HBP/SLBP functions during replication stress.