ASH2L Deficiency in Smooth Muscle Drives Pulmonary Vascular Remodeling

Abstract

Background: Histone H3 lysine 4 methylation is one of the most abundant epigenetic modifications, which has been recently linked to vascular remodeling in pulmonary hypertension (PH). SET1/MLL methyltransferase complexes comprise the main enzymes responsible for methylating H3 lysine 4, yet their roles in vascular remodeling and PH are not fully understood. We aim to assess the contribution of ASH2L, a core SET1/MLL family member, to the pathogenesis of PH.

Methods: Human pulmonary artery specimens and primary vascular cells, smooth muscle cell (SMC)-specific ASH2L-deficient mice, rats with SMC-specific ASH2L overexpression, mass spectrometry, immunoprecipitation, and chromatin immunoprecipitation were used to define the role of ASH2L in PH.

Results: Analysis of bulk RNA-sequencing data sets from human lung vessels identified ASH2L as the only differentially expressed SET1/MLL family member in PH compared with healthy controls. Decreased ASH2L expression in human pulmonary arteries correlated with the clinical severity of PH, which contrasted with elevated H3 lysine 4 methylation and was primarily localized to SMCs. Depletion of ASH2L promoted whereas its restoration ameliorated SMC proliferation and vascular remodeling in PH. Mechanistically, we revealed that ASH2L functioned independently of the canonical H3 lysine 4 trimethylation-based transcriptional activation, while it formed a protein complex with KLF5 and FBXW7, thereby accelerating the ubiquitin-proteasomal degradation of KLF5. NOTCH3 was discovered as a new downstream target of KLF5, and the loss of ASH2L promoted the recruitment of KLF5 to the NOTCH3 promoter, thus enhancing NOTCH3 expression. Pharmacological blockage of KLF5 attenuated PH in chronic hypoxia-exposed SMC-specific ASH2L-deficient mice and sugen/hypoxia-challenged rats.

Conclusions: This study demonstrated that ASH2L deficiency causatively affects SMC proliferation and lung vascular remodeling that is partially mediated through KLF5-dependent NOTCH3 transcription. Activating ASH2L or targeting KLF5 might represent potential therapeutic strategies for PH.

Keywords: RNA; cell proliferation; hypertension, pulmonary; muscle, smooth; vascular remodeling.