Linker histone H1 prevents R-loop accumulation and genome instability in heterochromatin

Aleix Bayona-Feliu,Jordi Bernués,Anna Casas-Lamesa,Fernando Azorín,Oscar Reina

doi:10.1038/s41467-017-00338-5

Abstract

Linker histone H1 is an important structural component of chromatin that stabilizes the nucleosome and compacts the nucleofilament into higher-order structures. The biology of histone H1 remains, however, poorly understood. Here we show that Drosophila histone H1 (dH1) prevents genome instability as indicated by the increased γH2Av (H2AvS137P) content and the high incidence of DNA breaks and sister-chromatid exchanges observed in dH1-depleted cells. Increased γH2Av occurs preferentially at heterochromatic elements, which are upregulated upon dH1 depletion, and is due to the abnormal accumulation of DNA:RNA hybrids (R-loops). R-loops accumulation is readily detectable in G1-phase, whereas γH2Av increases mainly during DNA replication. These defects induce JNK-mediated apoptosis and are specific of dH1 depletion since they are not observed when heterochromatin silencing is relieved by HP1a depletion. Altogether, our results suggest that histone H1 prevents R-loops-induced DNA damage in heterochromatin and unveil its essential contribution to maintenance of genome stability.

Highlights

Linker histone H1 is an important structural component of chromatin that stabilizes the nucleosome and compacts the nucleofilament into higher-order structures
Concomitant to increased γH2Av, Drosophila histone H1 (dH1)-depleted cells showed a high incidence of DNA breaks (DBs), as determined by the increased tail-moment observed in single-cell electrophoretic analyses performed under alkaline and neutral conditions to detect both single-stranded (SSBs) and double-stranded DNA breaks (DSBs) or only double-stranded breaks (DSBs), respectively (Fig. 1c)
DRIP-qPCR experiments showed that co-depletion of dH1 in HP1a-depleted cells restored R-loops accumulation in heterochromatin (Fig. 7c). These results indicate that, in HP1a-depleted cells, relief of heterochromatin silencing per se does not induce R-loops accumulation unless dH1 is depleted, suggesting that dH1 plays a specific role in preventing R-loops accumulation in heterochromatin

Summary

Introduction

Linker histone H1 is an important structural component of chromatin that stabilizes the nucleosome and compacts the nucleofilament into higher-order structures. We show that Drosophila histone H1 (dH1) prevents genome instability as indicated by the increased γH2Av (H2AvS137P) content and the high incidence of DNA breaks and sister-chromatid exchanges observed in dH1depleted cells. R-loops accumulation is readily detectable in G1-phase, whereas γH2Av increases mainly during DNA replication. These defects induce JNK-mediated apoptosis and are specific of dH1 depletion since they are not observed when heterochromatin silencing is relieved by HP1a depletion. Triple H1 variants knockout mice fail to compensate total H1 levels and die at early developmental stages[12] The reason for this lethality is not well understood. B WB analyses with αdH1, αγH2Av and αH4 of increasing amounts of extracts (lanes 1–3) prepared from siRNAdH1, siRNAlacZ and untreated cells. The p-value of siRNAdH1 respect to siRNAlacZ is indicated (***

Methods

Results

Conclusion