Epigenetic landscape of the H3K27me3 mark in macrophages transformed
byTheileria annulata
Authors: Sakura Takaya , Tajeri Shahin , Rchiad Zineb , Ansari Hifzur R. , Kaushik Abhinav , Mourier Tobias , Pain Arnab , Wassef Michel , Langsley Gordon 
Published: January 2025

Abstract

Significance statement
Tropical theileriosis is a major constraint to livestock production
worldwide. Infection withTheileria
annulatainduces the transformation of bovine leukocytes that
subsequently exhibit similarities to human leukemia.
Long-term in vitro culture of transformed
leukocytes attenuates their tumorigenic properties. The molecular changes
underlying this process are currently unclear. We focused on H3K27me3, a key
histone tail post-translational modification that regulates chromatin
structure and gene expression, and compared its genomic landscapes in
virulent, highly disseminating
macrophagesversusattenuated macrophages with
reduced dissemination potential that are used as a live vaccine against
tropical theileriosis. Western blot and ChIP-seq revealed higher global
levels of H3K27me3 and a striking broadening of its genomic distribution in
attenuated macrophages. To our surprise, this reconfiguration is not
associated with major changes in gene expression and attenuated cells appear
to be less responsive to PRC2 inhibition, suggesting that large-scale
broadening of H3K27me3 does not systematically translate into increased
PRC2-mediated silencing activity. Despite these changes, H3K27me3 maintains
the silencing of key tumor suppressor genes in both virulent and attenuated
macrophages. These findings shed light on the interplay between infection,
the evolution of oncogenic potential and reconfiguration of the chromatin
landscape following attenuation of virulence.

Theileria annulataandT.
parvaare obligate intracellular parasites that induce
bovine leukocyte transformation, leading to uncontrolled proliferation and
heightened dissemination of infected leukocytes, thus amplifying the
parasite and enhancing its transmission. Early passageT.
annulata-transformed macrophages are virulent, but
virulence decreases with long-term passage and infected macrophages become
attenuated for dissemination. Loss of dissemination is restored when
TGF-β2-mediated signaling is re-established in attenuated macrophages,
suggesting that genetic and/or epigenetic changes occurring within
macrophages contribute to the attenuated phenotype. To elucidate the
relationship between attenuation of dissemination and epigenetic
dysregulation ofT. annulata-transformed
macrophages, we focused on one of the important repressive histone marks,
tri-methylated lysine 27 of histone H3 (H3K27me3) catalysed by the Polycomb
Repressive Complex 2 (PRC2). We found that the global level of H3K27me3
ofT. annulata-infected macrophages is
increased in attenuated macrophages compared to virulent macrophages.
Chromatin immunoprecipitation sequencing (ChIP-seq) revealed that the
genomic distribution of H3K27me3 is heavily remodeled from virulent to
attenuated macrophages. In attenuated macrophages, many genes transition
from a focal peak of H3K27me3 around the transcription start site to larger
chromatin domains. But this transition is not paralleled by large-scale
transcriptional downregulation. In addition, RNAseq analysis following PRC2
inhibitor treatment reveals that fewer genes are derepressed in attenuated
macrophages than in virulent macrophages, suggesting that broader H3K27me3
profiles do not systematically translate into increased gene silencing
activity. Our findings shed light on the mechanisms underlying the
dysregulation of epigenetic modifications
inTheileria-induced leukocyte transformation
and contribute to a better understanding of the attenuation of tumor
dissemination.