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Fig. 6 | Epigenetics & Chromatin

Fig. 6

From: Spermatid-specific linker histone HILS1 is a poor condenser of DNA and chromatin and preferentially associates with LINE-1 elements

Fig. 6

Combinatorial histone PTMs identified in HILS1 containing chromatin in rat elongating spermatids. a Western blot analysis of the HILS1 pull-down fractions with antibodies against H4K5ac, H4K12ac, H3K9me3, H4K20me3, H3K4me3 and H3K27me3. Input (lane 1), IP using anti-HILS1 (lane 2), IgG (lane 3). No signal was observed with pre-immune IgG, and the heavy and light IgG chains are indicated by asterisk. Immunoprecipitation showed the specific enrichment of PTMs like H4 acetylation marks, H3K9me3 and H4K20me3 in HILS1 pull-down fraction, whereas methylation marks like H3K4me3 and H3K27me3 were completely absent in HILS1-associated chromatin. b Sequential ChIP assays of elongating/condensing spermatids using antibody against HILS1 followed by IP with H3K9me3/H4K20me3 antibody. qPCR analysis showed the specific enrichment of both the modifications in HILS1-associated loci (genomic positions are same as detailed in Fig. 6) in separate IP experiments and the negative control corresponds to a region in chr 8 depleted of HILS1. Data were quantified as % of input. Results are from three independent experiments, and error bars represent standard deviation. Asterisk shows comparison with negative control IgG for single IP, and # shows comparison with IgG+antibody2 for reChIP. ***P ≤ 0.001; **P ≤ 0.01; *P ≤ 0.05

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