{"title":"<i>METTL14</i>/<i>YTHDF1</i> mediates m6A modification of <i>PTBP1</i> to regulate PDGF-BB-induced airway smooth muscle cell function.","authors":"Canming Qiu, Zhenzhu Liao, Pingping Guo, Jun Liu","doi":"10.1080/01902148.2025.2546817","DOIUrl":null,"url":null,"abstract":"<p><strong>Background: </strong>Increased proliferation and migration of abnormal airway smooth muscle cells (ASMCs) are significantly associated with asthma. This study aimed to investigate the effects of methyltransferase-like 14 (<i>METTL14</i>), YTH domain-containing family Protein 1 (<i>YTHDF1</i>), and polypyrimidine tract-binding protein 1 (<i>PTBP1</i>) on platelet-derived growth factor-BB (PDGF-BB)-treated ASMCs.</p><p><strong>Methods: </strong>ASMCs were treated with PDGF-BB to mimic cell remodeling. A cell counting kit-8 (CCK-8) assay was performed to detect cell viability. Cell proliferation was detected by 5-Ethynyl-2'-deoxyuridine (EdU) assay. The migration and invasion of cells were measured by wound healing assay and transwell assay. Interleukin 1β (IL-1β) and tumor necrosis factor-α (TNF-α) were evaluated using ELISA kits. The oxidative stress markers reactive oxygen species (ROS) and malondialdehyde (MDA) levels were evaluated using corresponding kits. RT-qPCR and western blotting were utilized to assess mRNA and protein expression. The m6A level was determined using methylated RNA immunoprecipitation (MeRIP) assay. RNA Immunoprecipitation (RIP) assay was used to evaluate the binding of <i>METTL14</i> or <i>YTHDF1</i> to <i>PTBP1</i> mRNA. The binding of <i>METTL14</i> to <i>PTBP1</i> was quantified by dual-luciferase assay.</p><p><strong>Results: </strong>PDGF-BB treatment promoted ASMCs proliferation, migration, invasion, secretion of IL-1β and TNF-α, increased MDA and ROS levels, and promoted macrophage polarization. Knockdown of <i>PTBP1</i> attenuated PDGF-BB-induced proliferation, migration, invasion, inflammation, oxidative stress, and macrophage polarization in ASMCs. <i>METTL14</i>/<i>YTHDF1</i> facilitated the m6A methylation modification of <i>PTBP1</i>. Elevated <i>PTBP1</i> expression nullified the influence of increased <i>METTL14</i> expression on PDGF-BB-stimulated ASMCs. <i>METTL14</i> influenced the expression of nuclear factor kappa B (NF-κB) pathway-associated proteins <i>via PTBP1</i>.</p><p><strong>Conclusion: </strong>The m6A methylation of <i>PTBP1</i>, mediated by <i>METTL14</i>/<i>YTHDF1</i>, played a critical role in modulating the functional behavior of ASMCs induced by PDGF-BB during the progression of asthma.</p>","PeriodicalId":12206,"journal":{"name":"Experimental Lung Research","volume":"51 1","pages":"64-78"},"PeriodicalIF":1.8000,"publicationDate":"2025-01-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":"0","resultStr":null,"platform":"Semanticscholar","paperid":null,"PeriodicalName":"Experimental Lung Research","FirstCategoryId":"3","ListUrlMain":"https://doi.org/10.1080/01902148.2025.2546817","RegionNum":4,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":null,"EPubDate":"2025/8/23 0:00:00","PubModel":"Epub","JCR":"Q3","JCRName":"RESPIRATORY SYSTEM","Score":null,"Total":0}
引用次数: 0
Abstract
Background: Increased proliferation and migration of abnormal airway smooth muscle cells (ASMCs) are significantly associated with asthma. This study aimed to investigate the effects of methyltransferase-like 14 (METTL14), YTH domain-containing family Protein 1 (YTHDF1), and polypyrimidine tract-binding protein 1 (PTBP1) on platelet-derived growth factor-BB (PDGF-BB)-treated ASMCs.
Methods: ASMCs were treated with PDGF-BB to mimic cell remodeling. A cell counting kit-8 (CCK-8) assay was performed to detect cell viability. Cell proliferation was detected by 5-Ethynyl-2'-deoxyuridine (EdU) assay. The migration and invasion of cells were measured by wound healing assay and transwell assay. Interleukin 1β (IL-1β) and tumor necrosis factor-α (TNF-α) were evaluated using ELISA kits. The oxidative stress markers reactive oxygen species (ROS) and malondialdehyde (MDA) levels were evaluated using corresponding kits. RT-qPCR and western blotting were utilized to assess mRNA and protein expression. The m6A level was determined using methylated RNA immunoprecipitation (MeRIP) assay. RNA Immunoprecipitation (RIP) assay was used to evaluate the binding of METTL14 or YTHDF1 to PTBP1 mRNA. The binding of METTL14 to PTBP1 was quantified by dual-luciferase assay.
Results: PDGF-BB treatment promoted ASMCs proliferation, migration, invasion, secretion of IL-1β and TNF-α, increased MDA and ROS levels, and promoted macrophage polarization. Knockdown of PTBP1 attenuated PDGF-BB-induced proliferation, migration, invasion, inflammation, oxidative stress, and macrophage polarization in ASMCs. METTL14/YTHDF1 facilitated the m6A methylation modification of PTBP1. Elevated PTBP1 expression nullified the influence of increased METTL14 expression on PDGF-BB-stimulated ASMCs. METTL14 influenced the expression of nuclear factor kappa B (NF-κB) pathway-associated proteins via PTBP1.
Conclusion: The m6A methylation of PTBP1, mediated by METTL14/YTHDF1, played a critical role in modulating the functional behavior of ASMCs induced by PDGF-BB during the progression of asthma.
期刊介绍：
Experimental Lung Research publishes original articles in all fields of respiratory tract anatomy, biology, developmental biology, toxicology, and pathology. Emphasis is placed on investigations concerned with molecular, biochemical, and cellular mechanisms of normal function, pathogenesis, and responses to injury. The journal publishes reports on important methodological advances on new experimental modes. Also published are invited reviews on important and timely research advances, as well as proceedings of specialized symposia.
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