METTL3 mediates SOX5 m6A methylation in bronchial epithelial cells to attenuate Th2 cell differentiation in T2 asthma

Zhifeng Chen; Yulin Shang; Xiufeng Zhang; Wentao Duan; Jianmin Li; Liming Zhu; Libing Ma; Xudong Xiang; Jingsi Jia; Xiaoying Ji; Subo Gong

doi:10.1016/j.heliyon.2024.e28884

METTL3 mediates SOX5 m6A methylation in bronchial epithelial cells to attenuate Th2 cell differentiation in T2 asthma

Heliyon. 2024 Apr 1;10(7):e28884. doi: 10.1016/j.heliyon.2024.e28884. eCollection 2024 Apr 15.

Authors

Zhifeng Chen¹, Yulin Shang², Xiufeng Zhang³, Wentao Duan⁴, Jianmin Li⁴, Liming Zhu⁴, Libing Ma⁵, Xudong Xiang⁶, Jingsi Jia⁶, Xiaoying Ji⁷, Subo Gong⁸

Affiliations

¹ Department of Respiratory and Critical Care Medicine, The Second Xiangya Hospital, Central South University, 139 Middle Renmin Road, Changsha, Hunan, 410011, China.
² Ophthalmology and Otorhinolaryngology, Zigui County Traditional Chinese Medicine Hospital, 30 Pinghu Avenue, Zigui, Hubei, 443600, China.
³ Department of Respiratory and Critical Care Medicine, The Second Affiliated Hospital of Hainan Medical University, 48 Pak Shui Tong Road, Haikou, Hainan, 570000, China.
⁴ Department of Respiratory and Critical Care Medicine, Hunan Provincial People's Hospital, 61 West Jiefang Road, Changsha, Hunan, 410005, China.
⁵ Department of Respiratory and Critical Care Medicine, The Affiliated Hospital of Guilin Medical University, 15 Le Qun Road, Guilin, Guangxi, 541001, China.
⁶ Department of Emergency, The Second Xiangya Hospital, Central South University, 139 Middle Renmin Road, Changsha, Hunan, 410011, China.
⁷ Department of Respiratory and Critical Care Medicine, The Affiliated Hospital of Guizhou Medical University, 28 Guiyi Street, Guiyang, Guizhou, 550004, China.
⁸ Department of Geriatrics, The Second Xiangya Hospital, Central South University, 139 Middle Renmin Road, Changsha, Hunan, 410011, China.

Abstract

Objective: Asthma, a chronic inflammatory disease in which type 2 T helper cells (Th2) play a causative role in the development of T2 asthma. N6-methyladenosine (m6A) modification, an mRNA modification, and methyltransferase-like 3 (METTL3) is involved in the development of T2 asthma by inhibiting Th2 cell differentiation. Sex determining region Y-box protein 5 (SOX5) is involved in regulating T cell differentiation, but its role in T2 asthma was unclear. The objective of this study was to explore the role of METTL3 and SOX5 in T2 asthma and whether there is an interaction between the two.

Materials and methods: Adults diagnosed with T2 asthma (n = 14) underwent clinical information collection and pulmonary function tests. In vivo and in vitro T2 asthma models were established using female C57BL/6 mice and human bronchial epithelial cells (HBE). The expressions of METTL3 and SOX5 were detected by Western blot and qRT-PCR and Western blot. Th2 cell differentiation was determined by flow cytometry and IL-4 level was detected by ELISA. m6A methylation level was determined by m6A quantitative assay. The relationship between METTL3 expression and clinical parameters was determined by Spearman rank correlation analysis. The function of METTL3 and SOX5 genes in asthma was investigated in vitro and in vivo. The RNA immunoprecipitation assay detected the specific interaction between METTL3 and SOX5.

Results: Patients with T2 asthma displayed lower METTL3 levels compared to healthy controls. Within this group, a negative correlation was observed between METTL3 and Th2 cells, while a positive correlation was noted between METTL3 and clinical parameters as well as Th1 cells. In both in vitro and in vivo models representing T2 asthma, METTL3 levels decreased significantly, while SOX5 levels showed the opposite trend. Overexpression of METTL3 gene in HBE cells significantly inhibited Th2 cell differentiation and increased m6A methylation activity. From a mechanism perspective, low METTL3 negatively regulates SOX5 expression through m6A modification dependence, while high SOX5 expression is positively associated with T2 asthma severity. Cell transfection experiments confirmed that METTL3 regulates Th2 cell differentiation and IL-4 release through SOX5.

Conclusions: Overall, our results indicate that METTL3 alleviates Th2 cell differentiation in T2 asthma by modulating the m6A methylation activity of SOX5 in bronchial epithelial cells. This mechanism could potentially serve as a target for the prevention and management of T2 asthma.

Keywords: HBE cells; METTL3; SOX5; T2 asthma; Th2; m6A.