Histone H1 limits DNA methylation inNeurospora crassa

Abstract

Histone H1 variants, known as linker histones, are essential chromatin components in higher eukaryotes, yet compared to the core histones relatively little is known about their in vivo functions. The filamentous fungus Neurospora crassa encodes a single H1 protein that is not essential for viability. To investigate the role of N. crassa H1, we constructed a functional FLAG-tagged H1 fusion protein and performed genomic and molecular analyses. Cell fractionation experiments showed that H1-3XFLAG is a chromatin binding protein. Chromatin-immunoprecipitation combined with sequencing (ChIP-seq) revealed that H1-3XFLAG is globally enriched throughout the genome with a subtle preference for promoters of expressed genes. In mammals, the stoichiometry of H1 impacts nucleosome repeat length. To determine if H1 impacts nucleosome occupancy or nucleosome positioning in N. crassa, we performed micrococcal nuclease digestion in the wild-type and the DhH1 strain followed by sequencing (MNase-seq). Deletion of hH1 did not significantly impact nucleosome positioning or nucleosome occupancy. Analysis of DNA methylation by whole-genome bisulfite sequencing (MethylC-seq) revealed a modest but global increase in DNA methylation in the DhH1 mutant. Together, these data suggest that H1 acts as a nonspecific chromatin binding protein that can limit accessibility of the DNA methylation machinery in N. crassa.

Centromere evolution in the fungal genusVerticillium

“…2a; Fig. S2), similar to the previously reported levels for Aspergillus flavus (46) and lower than for N. crassa (47). Nevertheless, repetitive elements and centromeres show significantly higher DNA methylation levels in all contexts when compared with genes ( Fig.…”

DNA Methylation and Histone H1 Jointly Repress Transposable Elements and Aberrant Intragenic Transcripts

“…There is now extensive evidence that H1 modulates DNA methylation pathways. Loss of H1 causes genome-wide hypermethylation in ascomycete fungi (Barra et al, 2000;Seymour et al, 2016) and reduces methylation at specific loci in mouse (Fan et al, 2005;Geeven et al, 2015;Maclean et al, 2011;Yang et al, 2013). In Arabidopsis, loss of H1 reduces methylation of euchromatic TEs and increases methylation of heterochromatic elements in all sequence contexts (Rutowicz et al, 2015;Zemach et al, 2013).…”

Chromatin accessibility profiling in Neurospora crassa reveals molecular features associated with accessible and inaccessible chromatin

“…Libraries were cleaned using SeraPure beads (Fisher Cat. # 09-981-123) before submission for sequencing as described in [80,81]. Libraries were sequenced on an Illumina NextSeq 500 instrument by Georgia Genomics Facility at University of Georgia.…”

“…ChIP-seq was performed as described in [52,80,81] with the following antibodies: H3K27me2/3 (Active Motif Cat# 39535, Lot# 1671401)2; H3K4me1 (Abcam Cat# ab9885); H3K4me2 (Active Motif Cat# 29679, Lot# 31713007); H3K4me3 (Abcam Cat# ab1012); H3K36me3 (Abcam Cat# ab9050, Lot# GR300388-1); H3K9me3 (Active Motif Cat# 39161, Lot# 14418003); H3K27ac; FLAG (Sigma Cat# F1804, Lot# SLBS3530V). Libraries were created and analyzed as described in [81].…”