Characterization of maternal histone mRNAs

Abstract

[eng] In eukaryotes, histone proteins bind and pack genomic DNA into chromatin. The basic structural subunit of chromatin is the nucleosome, which is formed by the interaction of an octamer of the core histone proteins H2A, H2B, H3 and H4 with 147 bp of DNA. In addition, linker histones H1 bind to the nucleosome core particle at the entry/exit site of nucleosomal DNA and interact with the internucleosomal linker DNA. According to their pattern of expression during the cell cycle, histones are classified into replication dependent (RD), whose expression is tightly linked to DNA replication, and replication independent (RI). RD histones include the four canonical core histones and most linker histones H1 and are involved in packaging the newly generated DNA during replication. On the other hand, histone variants are generally RI. RD histone mRNAs are distinct from every other mRNA in the cell since they are not polyadenylated. Instead, they have a conserved sequence forming a stem-loop at the 3’UTR, which is recognized by the Stem-Loop Binding Protein (SLBP). SLBP is responsible for processing, stabilization, and translation of RD histone mRNAs, as well as for cell cycle coordination of their expression. In contrast, RI histone mRNAs are polyadenylated. In metazoans, early embryogenesis usually involves rapid nuclear divisions in the absence of zygotic expression. The number and speed of these divisions are generally higher in species with external development. In Drosophila, there are 13 syncytial nuclear divisions before zygotic genome activation (ZGA). During these early divisions, dSLBP is absent, and packaging of the rapidly generated DNA relies on maternal histones. Core RD histones are maternally deposited as both proteins and mRNAs. Instead, we found that the single RD linker histone of Drosophila, dH1, is deposited only as mRNAs. In addition to dH1, Drosophila encodes for a germline-specific linker histone dBigH1 variant, which is maternally deposited as both protein and mRNA. In this study, we show that, in Drosophila, maternally deposited RD histone mRNAs are polyadenylated and have a truncated 3’UTRs that disrupts the characteristic 3’ stem-loop structure. We also show that these

unusual RD histone transcripts are generated during oogenesis, in stage 10 egg chambers, through an alternative processing that surprisingly requires dSLBP and involves cytoplasmic polyadenylation. Polyadenylation of maternal RD histone mRNAs does not appear to be restricted to Drosophila since it is also observed in Xenopus, which also undergoes external development. Intriguingly, we found that maternal RD histone mRNAs remain largely untranslated during early embryogenesis. In particular, despite being maternally deposited as mRNAs, expression of the single RD linker histone of Drosophila, dH1, is not detected during the early embryo divisions. Instead, at these stages, the germline-specific linker histone dBigH1 variant is expressed. dH1 expression starts at nuclear division 6 and increases progressively during ZGA, fully replacing dBigH1 at cellularization. Notably, we found that loss of dBigH1 in homozygous null bigH1 mutant embryos induces the early expression of dH1 from the first nuclear division, which is concomitant to increased translation of maternal dH1 mRNAs. Interestingly, homozygous null bigH1 mutant embryos progress normally through development. These results unveil the compensatory expression of dH1 in the absence of dBigH1, supporting functional redundancy of dBigH1 and dH1 during early embryogenesis. Finally, we show that translation of the rest of maternal RD histone transcripts is also increased in the absence of dBigH1. Altogether, these results suggest that translation of the maternal pool of RD histone transcripts serves as a backup mechanism that is induced in response to reduced supply of maternal histone proteins.