DOI: 10.5281/zenodo.4139402
title: Transcriptome analysis of the effect of over-expressing H2A.J mutants in proliferating WI38 fibroblasts for the paper entitled: The H2A.J histone variant contributes to Interferon-Stimulated Gene expression in senescence by its weak interaction with H1 and the derepression of repeated DNA sequences

description: Abstract for overall study: The histone variant H2A.J was previously shown to accumulate in senescent human fibroblasts with persistent DNA damage to promote inflammatory gene expression, but its mechanism of action was unknown. We show that H2A.J accumulation contributes to weakening the association of histone H1 to chromatin and increasing its turnover. Decreased H1 in senescence is correlated with increased expression of some repeated DNA sequences, increased expression of STAT/IRF transcription factors, and transcriptional activation of Interferon-Stimulated Genes (ISGs). The H2A.J-specific Val-11 moderates the transcriptional activity of H2A.J, and H2A.J-specific Ser-123 can be phosphorylated in response to DNA damage with potentiation of its transcriptional activity by the phospho-mimetic S123E mutation. Our work demonstrates the functional importance of H2A.J-specific residues and potential mechanisms for its function in promoting inflammatory gene expression in senescence. Specific description for this dataset: H2A.J differs from canonical H2A only by a valine at position 11 instead of alanine, and the 7 C-terminal amino acids containing a potential minimal phosphorylation site SQ for DNA-damage response kinases. To test the functional importance of these H2A.J-specific sequences, we mutated Val-11 to Ala as is found in all canonical H2A sequences, and we mutated Ser-123 to either Glu to mimic a phospho-serine residue or to Ala to prevent phosphorylation. We also substituted the C-terminus of H2A.J with the C-terminus of H2A. These mutants, WT-H2A.J and canonical H2A-type1 were ectopically expressed in proliferating fibroblasts, and their microarray transcriptomes were compared to that of proliferating and senescent fibroblasts without ectopic histone expression. Genome-wide transcriptome analysis indicated that senescent fibroblasts clustered distinctly from proliferating fibroblasts, and proliferating fibroblasts expressing the H2A.J-V11A and H2A.J-S123E mutants clustered distinctly from fibroblasts expressing the other H2A.J mutants, WT-H2A.J, and H2A. Hallmark gene set enrichment analysis of the transcriptomes of fibroblasts expressing H2A.J-V11A or H2A.J-S123E versus control proliferating fibroblasts indicated that they showed the same highly significant enrichment for the Epithelial-Mesenchyme Transition, TNF-Alpha Signaling Via NF-kB, and Inflammatory Response gene sets. Notable inflammatory genes including IL1A, IL1B, IL6, CXCL8, and CCL2 are contained in these gene sets and are often induced in senescence as part of the senescence-associated secretory phenotype. Heat maps showed that the H2A.J-V11A and H2A.J-S123E mutants were particularly apt at activating the expression of these inflammatory genes in proliferating fibroblasts
description: BioRxiv preprint: doi: https://doi.org/10.1101/2020.10.29.361204
description: {"references": ["BioRxiv preprint: doi: https://doi.org/10.1101/2020.10.29.361204"]}