Ezh2 Shapes T Cell Plasticity to Drive Atherosclerosis
Authors: Bonfiglio Cecilia Assunta , Lacy Michael , Triantafyllidou Vasiliki , Farina Floriana Maria , Janjic Aleksandar , Nitz Katrin , Wu Yuting , Bazioti Venetia , Avcilar-Kucukgoze Irem , Marques Yonara Freire Soares , Joppich Markus , Kumkum Mahadia , Röß Katja , Venkatasubramani Anuroop Venkateswaran , Imhof Axel , Enard Wolfgang , Maegdefessel Lars , de Winther Menno , Weber Christian , Santovito Donato , Lutgens Esther , Atzler Dorothee 
Published: February 2025

BACKGROUND:

The activation and polarization of T cells play a crucial role in atherosclerosis and dictate athero-inflammation. The epigenetic enzyme EZH2 (enhancer of zeste homolog 2) mediates the H3K27me3 (trimethylation of histone H3 lysine 27) and is pivotal in controlling T cell responses.

METHODS:

To detail the role of T cell EZH2 in atherosclerosis, we used human carotid endarterectomy specimens to reveal plaque expression and geography of EZH2. Atherosclerosis-prone
Apoe
(apolipoprotein E)–deficient mice with CD (cluster of differentiation) 4
+
or CD8
+
T cell–specific
Ezh2
deletion (Ezh2
cd4
-knockout [KO], Ezh2
cd8
-KO) were analyzed to unravel the role of T cell Ezh2 in atherosclerosis and T cell–associated immune status.

RESULTS:

EZH2
expression is elevated in advanced human atherosclerotic plaques and primarily expressed in the T cell nucleus, suggesting the importance of canonical EZH2 function in atherosclerosis. Ezh2
cd4
-KO, but not Ezh2
cd8
-KO, mice showed reduced atherosclerosis with fewer advanced plaques, which contained less collagen and macrophages, indicating that Ezh2 in CD4
+
T cells drives atherosclerosis. In-depth analysis of CD4
+
T cells of Ezh2
cd4
-KO mice revealed that absence of Ezh2 results in a type 2 immune response with increased Il-4 (interleukin 4) gene and protein expression in the aorta and lymphoid organs. In vitro,
Ezh2
-deficient T cells polarized macrophages toward an anti-inflammatory phenotype. Single-cell RNA-sequencing of splenic T cells revealed that
Ezh2
deficiency reduced naive, Ccl5
+
(C-C motif chemokine ligand 5) and regulatory T cell populations and increased the frequencies of memory T cells and invariant natural killer T (iNKT) cells. Flow cytometric analysis identified a shift toward Th2 (type 2 T helper) effector CD4
+
T cells in Ezh2
cd4
-KO mice and confirmed a profound increase in splenic iNKT cells with increased expression of Plzf (promyelocytic leukemia zinc finger), which is the characteristic marker of the iNKT2 subset. Likewise,
Zbtb16
([zinc finger and BTB domain containing 16], a Plzf-encoding gene) transcripts were elevated in the aorta of Ezh2
cd4
-KO mice, suggesting an accumulation of iNKT2 cells in the plaque. H3K27me3–chromatin immunoprecipitation followed by quantitative polymerase chain reaction showed that T cell–Ezh2 regulates the transcription of the
Il-4
and
Zbtb16
genes.

CONCLUSIONS:

Our study uncovers the importance of T cell EZH2 in human and mouse atherosclerosis. Inhibition of Ezh2 in CD4
+
T cells drives type 2 immune responses, resulting in an accumulation of iNKT2 and Th2 cells, memory T cells and anti-inflammatory macrophages that limit the progression of atherosclerosis.