RNA结合蛋白ZFP36L1和ZFP36L2在人类和哮喘小鼠模型的气道上皮中表达失调
2023/11/23
摘要
背景：哮喘是最常见的慢性气道炎症性疾病。气道上皮是该病的关键驱动因素，许多研究已经确认健康和哮喘之间在mRNA表达上存在基因组范围的差异。然而，这些差异的潜在分子机制仍然知之甚少。人类TTP家族由ZFP36、ZFP36L1和ZFP36L2组成，在免疫调节中发挥着关键作用，通过决定编码炎症介质的众多mRNAs的稳定性和翻译来实现。我们调查了RNA结合蛋白（RBPs）TTP家族在哮喘中的表达和可能的作用，但对其在哮喘中的作用知之甚少。
方法：我们分析了多个公开的哮喘数据集（包括单细胞RNA测序）中ZFP36、ZFP36L1和ZFP36L2 mRNA的水平。我们还对这些RBPs已知靶点在哮喘中的表达进行了研究。我们在小鼠哮喘模型的精确切片中评估了Zfp36l1和Zfp36l2的肺mRNA表达和细胞定位。最后，我们在重症哮喘患者的原代支气管上皮中确定了ZFP36L1和ZFP36L2的表达，并进行了回复实验。
结果：我们发现在不同的队列中（5例健康对照与8例重度哮喘；36例中度哮喘对比37例吸入类固醇治疗的重度哮喘；26例口服皮质激素治疗的重度哮喘），ZFP36L1和ZFP36L2的mRNA水平在重度哮喘患者的气道上皮中显著下调。通过整合多个数据集，我们推断这些RBPs可能靶向的mRNAs在重度哮喘中增加。在哮喘小鼠模型中，Zfp36l1在肺中下调，通过对离体肺片的双抗体免疫染色显示，在急性哮喘小鼠模型的气道上皮中，Zfp36l1/l2的核定位增加，而在慢性模型中进一步增强。人类支气管活检的免疫染色显示，与轻度相比，重度哮喘患者气道上皮细胞对ZFP36L1的染色减少，而ZFP36L2上调。在重度哮喘患者的原代支气管上皮细胞中恢复ZFP36L1和ZFP36L2水平降低了IL6、IL8和CSF2的mRNA表达。
结论：我们认为，ZFP36L1/L2 水平的失调及其亚细胞错定位是导致哮喘患者 mRNA 表达和细胞质命运发生变化的原因。
The RNA binding proteins ZFP36L1 and ZFP36L2 are dysregulated in airway epithelium in human and a murine model of asthma
Rynne J, Ortiz-Zapater E, Bagley DC, Zanin O, Doherty G, Kanabar V, Ward J, Jackson DJ, Parsons M, Rosenblatt J, Adcock IM, Martinez-Nunez RT.
Abstract
Background:Asthma is the most common chronic inflammatory disease of the airways. The airway epithelium is a key driver of the disease, and numerous studies have established genome-wide differences in mRNA expression between health and asthma. However, the underlying molecular mechanisms for such differences remain poorly understood. The human TTP family is comprised of ZFP36, ZFP36L1 and ZFP36L2, and has essential roles in immune regulation by determining the stability and translation of myriad mRNAs encoding for inflammatory mediators. We investigated the expression and possible role of the tristetraprolin (TTP) family of RNA binding proteins (RBPs), poorly understood in asthma.
Methods:We analysed the levels of ZFP36, ZFP36L1 and ZFP36L2 mRNA in several publicly available asthma datasets, including single cell RNA-sequencing. We also interrogated the expression of known targets of these RBPs in asthma. We assessed the lung mRNA expression and cellular localization of Zfp36l1 and Zfp36l2 in precision cut lung slices in murine asthma models. Finally, we determined the expression in airway epithelium of ZFP36L1 and ZFP36L2 in human bronchial biopsies and performed rescue experiments in primary bronchial epithelium from patients with severe asthma.
Results:We found ZFP36L1 and ZFP36L2 mRNA levels significantly downregulated in the airway epithelium of patients with very severe asthma in different cohorts (5 healthy vs. 8 severe asthma; 36 moderate asthma vs. 37 severe asthma on inhaled steroids vs. 26 severe asthma on oral corticoids). Integrating several datasets allowed us to infer that mRNAs potentially targeted by these RBPs are increased in severe asthma. Zfp36l1 was downregulated in the lung of a mouse model of asthma, and immunostaining of ex vivo lung slices with a dual antibody demonstrated that Zfp36l1/l2 nuclear localization was increased in the airway epithelium of an acute asthma mouse model, which was further enhanced in a chronic model. Immunostaining of human bronchial biopsies showed that airway epithelial cell staining of ZFP36L1 was decreased in severe asthma as compared with mild, while ZFP36L2 was upregulated. Restoring the levels of ZFP36L1 and ZFP36L2 in primary bronchial epithelial cells from patients with severe asthma decreased the mRNA expression of IL6, IL8 and CSF2.
Conclusion:We propose that the dysregulation of ZFP36L1/L2 levels as well as their subcellular mislocalization contributes to changes in mRNA expression and cytoplasmic fate in asthma.
背景：哮喘是最常见的慢性气道炎症性疾病。气道上皮是该病的关键驱动因素，许多研究已经确认健康和哮喘之间在mRNA表达上存在基因组范围的差异。然而，这些差异的潜在分子机制仍然知之甚少。人类TTP家族由ZFP36、ZFP36L1和ZFP36L2组成，在免疫调节中发挥着关键作用，通过决定编码炎症介质的众多mRNAs的稳定性和翻译来实现。我们调查了RNA结合蛋白（RBPs）TTP家族在哮喘中的表达和可能的作用，但对其在哮喘中的作用知之甚少。
方法：我们分析了多个公开的哮喘数据集（包括单细胞RNA测序）中ZFP36、ZFP36L1和ZFP36L2 mRNA的水平。我们还对这些RBPs已知靶点在哮喘中的表达进行了研究。我们在小鼠哮喘模型的精确切片中评估了Zfp36l1和Zfp36l2的肺mRNA表达和细胞定位。最后，我们在重症哮喘患者的原代支气管上皮中确定了ZFP36L1和ZFP36L2的表达，并进行了回复实验。
结果：我们发现在不同的队列中（5例健康对照与8例重度哮喘；36例中度哮喘对比37例吸入类固醇治疗的重度哮喘；26例口服皮质激素治疗的重度哮喘），ZFP36L1和ZFP36L2的mRNA水平在重度哮喘患者的气道上皮中显著下调。通过整合多个数据集，我们推断这些RBPs可能靶向的mRNAs在重度哮喘中增加。在哮喘小鼠模型中，Zfp36l1在肺中下调，通过对离体肺片的双抗体免疫染色显示，在急性哮喘小鼠模型的气道上皮中，Zfp36l1/l2的核定位增加，而在慢性模型中进一步增强。人类支气管活检的免疫染色显示，与轻度相比，重度哮喘患者气道上皮细胞对ZFP36L1的染色减少，而ZFP36L2上调。在重度哮喘患者的原代支气管上皮细胞中恢复ZFP36L1和ZFP36L2水平降低了IL6、IL8和CSF2的mRNA表达。
结论：我们认为，ZFP36L1/L2 水平的失调及其亚细胞错定位是导致哮喘患者 mRNA 表达和细胞质命运发生变化的原因。
(中日友好医院呼吸与危重症医学科 沈焜路 摘译 林江涛 审校)
(Front Cell Dev Biol. 2023 Oct 19. DOI: 10.3389/fcell.2023.1241008)
(Front Cell Dev Biol. 2023 Oct 19. DOI: 10.3389/fcell.2023.1241008)
The RNA binding proteins ZFP36L1 and ZFP36L2 are dysregulated in airway epithelium in human and a murine model of asthma
Rynne J, Ortiz-Zapater E, Bagley DC, Zanin O, Doherty G, Kanabar V, Ward J, Jackson DJ, Parsons M, Rosenblatt J, Adcock IM, Martinez-Nunez RT.
Abstract
Background:Asthma is the most common chronic inflammatory disease of the airways. The airway epithelium is a key driver of the disease, and numerous studies have established genome-wide differences in mRNA expression between health and asthma. However, the underlying molecular mechanisms for such differences remain poorly understood. The human TTP family is comprised of ZFP36, ZFP36L1 and ZFP36L2, and has essential roles in immune regulation by determining the stability and translation of myriad mRNAs encoding for inflammatory mediators. We investigated the expression and possible role of the tristetraprolin (TTP) family of RNA binding proteins (RBPs), poorly understood in asthma.
Methods:We analysed the levels of ZFP36, ZFP36L1 and ZFP36L2 mRNA in several publicly available asthma datasets, including single cell RNA-sequencing. We also interrogated the expression of known targets of these RBPs in asthma. We assessed the lung mRNA expression and cellular localization of Zfp36l1 and Zfp36l2 in precision cut lung slices in murine asthma models. Finally, we determined the expression in airway epithelium of ZFP36L1 and ZFP36L2 in human bronchial biopsies and performed rescue experiments in primary bronchial epithelium from patients with severe asthma.
Results:We found ZFP36L1 and ZFP36L2 mRNA levels significantly downregulated in the airway epithelium of patients with very severe asthma in different cohorts (5 healthy vs. 8 severe asthma; 36 moderate asthma vs. 37 severe asthma on inhaled steroids vs. 26 severe asthma on oral corticoids). Integrating several datasets allowed us to infer that mRNAs potentially targeted by these RBPs are increased in severe asthma. Zfp36l1 was downregulated in the lung of a mouse model of asthma, and immunostaining of ex vivo lung slices with a dual antibody demonstrated that Zfp36l1/l2 nuclear localization was increased in the airway epithelium of an acute asthma mouse model, which was further enhanced in a chronic model. Immunostaining of human bronchial biopsies showed that airway epithelial cell staining of ZFP36L1 was decreased in severe asthma as compared with mild, while ZFP36L2 was upregulated. Restoring the levels of ZFP36L1 and ZFP36L2 in primary bronchial epithelial cells from patients with severe asthma decreased the mRNA expression of IL6, IL8 and CSF2.
Conclusion:We propose that the dysregulation of ZFP36L1/L2 levels as well as their subcellular mislocalization contributes to changes in mRNA expression and cytoplasmic fate in asthma.
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