Identification of a distal RXFP1 gene enhancer with differential activity in fibrotic lung fibroblasts involving AP-1.
Identification of a distal RXFP1 gene enhancer with differential activity in fibrotic lung fibroblasts involving AP-1.
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DOI：
10.1371/journal.pone.0254466
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发表时间：
2021
期刊：
影响因子：
3.7
通讯作者：
Zhang Y
中科院分区：
文献类型：
--
作者：
Chen TY;Li X;Goobie GC;Hung CH;Hung TK;Hamilton K;Bahudhanapati H;Tan J;Kass DJ;Zhang Y
松弛素/胰岛素样家族肽受体1（RXFP 1）介导松弛素的抗纤维化作用，并在纤维化间质性肺病（fILD）（包括特发性肺纤维化（IPF）和系统性硬化症（SSc））患者的肺和皮肤中表达减少。这可能解释了基于松弛素的抗纤维化治疗在SSc中的失败，但控制RXFP 1表达的调节机制在很大程度上仍然未知。本研究旨在确定RXFP 1的调控元件，可能在纤维化成纤维细胞中发挥差异作用。我们使用荧光素酶报告系统鉴定并评估了肺成纤维细胞中RXFP 1的远端调控区。使用连续缺失，上调pGL 3-启动子活性的增强子定位于距远端转录起始位点（TSS）-584至-242bp之间的远端区域。该增强子在IPF和SSc肺成纤维细胞中表现出降低的活性。生物信息学分析确定了两个集群的激活蛋白1（AP-1）转录因子结合位点的增强子。结合位点的定点诱变证实仅一个簇降低活性（相对于远端TSS为-358至-353）。在肺成纤维细胞中共表达FOS进一步增加增强子活性。使用从肺成纤维细胞分离的核蛋白，用跨越功能性AP-1位点的标记探针在体外形成复合物，证实了特异性DNA/蛋白质复合物的形成。应用抗JUN和FOS的抗体导致了复杂的改变，而抗JUNB和FOSL 1的抗体则没有。5对对照和IPF肺成纤维细胞中AP-1结合的分析在对照成纤维细胞中检测到更频繁的阳性结合。IPF肺组织中JUN和FOS的表达降低，并与RXFP 1的表达呈正相关。总之，我们确定了一个远端增强子RXFP 1与差异活性纤维化肺成纤维细胞涉及AP-1转录因子。我们的研究提供了对fILD中RXFP 1下调的深入了解，并可能支持重新评估基于松弛素的治疗方法以及RXFP 1转录上调的努力。
Relaxin/insulin-like family peptide receptor 1 (RXFP1) mediates relaxin’s antifibrotic effects and has reduced expression in the lung and skin of patients with fibrotic interstitial lung disease (fILD) including idiopathic pulmonary fibrosis (IPF) and systemic sclerosis (SSc). This may explain the failure of relaxin-based anti-fibrotic treatments in SSc, but the regulatory mechanisms controlling RXFP1 expression remain largely unknown. This study aimed to identify regulatory elements of RXFP1 that may function differentially in fibrotic fibroblasts. We identified and evaluated a distal regulatory region of RXFP1 in lung fibroblasts using a luciferase reporter system. Using serial deletions, an enhancer upregulating pGL3-promoter activity was localized to the distal region between -584 to -242bp from the distal transcription start site (TSS). This enhancer exhibited reduced activity in IPF and SSc lung fibroblasts. Bioinformatic analysis identified two clusters of activator protein 1 (AP-1) transcription factor binding sites within the enhancer. Site-directed mutagenesis of the binding sites confirmed that only one cluster reduced activity (-358 to -353 relative to distal TSS). Co-expression of FOS in lung fibroblasts further increased enhancer activity. In vitro complex formation with a labeled probe spanning the functional AP-1 site using nuclear proteins isolated from lung fibroblasts confirmed a specific DNA/protein complex formation. Application of antibodies against JUN and FOS resulted in the complex alteration, while antibodies to JUNB and FOSL1 did not. Analysis of AP-1 binding in 5 pairs of control and IPF lung fibroblasts detected positive binding more frequently in control fibroblasts. Expression of JUN and FOS was reduced and correlated positively with RXFP1 expression in IPF lungs. In conclusion, we identified a distal enhancer of RXFP1 with differential activity in fibrotic lung fibroblasts involving AP-1 transcription factors. Our study provides insight into RXFP1 downregulation in fILD and may support efforts to reevaluate relaxin-based therapeutics alongside upregulation of RXFP1 transcription.
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DOI：
10.1124/jpet.110.170977
发表时间：
2010-12-01
影响因子：
3.5
作者：
Pini, Alessandro;Shemesh, Ronen;Rotman, Galit
通讯作者：
Rotman, Galit
影响因子：
4.1
作者：
Ahmad, Nisar;Wang, Wei;Nair, Remi;Kapila, Sunil
通讯作者：
Kapila, Sunil
影响因子：
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作者：
Khanna, Dinesh;Clements, Philip J.;Seibold, James R.
通讯作者：
Seibold, James R.
影响因子：
5.3
作者：
Leppä, S;Eriksson, M;Bohmann, D
通讯作者：
Bohmann, D
DOI：
10.1016/s0140-6736(18)32335-3
发表时间：
2018-11-10
期刊：
Lancet (London, England)
影响因子：
--
作者：
GBD 2017 DALYs and HALE Collaborators
通讯作者：
GBD 2017 DALYs and HALE Collaborators