狼疮皮肤中 Hippo 通路对光敏感性的调节。
Regulation of Photosensitivity by the Hippo Pathway in Lupus Skin.
机构信息
Department of Dermatology, University of Michigan, Ann Arbor.
Division of Rheumatology and Graduate Program in Immunology, University of Michigan, Ann Arbor, and Departments of Pediatrics, Medicine, and Immunology, and Lupus Center of Excellence, University of Pittsburgh, Pittsburgh, Pennsylvania.
出版信息
Arthritis Rheumatol. 2023 Jul;75(7):1216-1228. doi: 10.1002/art.42460. Epub 2023 Apr 18.
OBJECTIVE
Photosensitivity is one of the most common manifestations of systemic lupus erythematosus (SLE), yet its pathogenesis is not well understood. The normal-appearing epidermis of patients with SLE exhibits increased ultraviolet B (UVB)-driven cell death that persists in cell culture. Here, we investigated the role of epigenetic modification and Hippo signaling in enhanced UVB-induced apoptosis seen in SLE keratinocytes.
METHODS
We analyzed DNA methylation in cultured keratinocytes from SLE patients compared to keratinocytes from healthy controls (n = 6/group). Protein expression was validated in cultured keratinocytes using immunoblotting and immunofluorescence. An immortalized keratinocyte line overexpressing WWC1 was generated via lentiviral vector. WWC1-driven changes were inhibited using a large tumor suppressor kinase 1/2 (LATS1/2) inhibitor (TRULI) and small interfering RNA (siRNA). The interaction between the Yes-associated protein (YAP) and the transcriptional enhancer associate domain (TEAD) was inhibited by overexpression of an N/TERT cell line expressing a tetracycline-inducible green fluorescent protein-tagged protein that inhibits YAP-TEAD binding (TEADi). Apoptosis was assessed using cleaved caspase 3/7 and TUNEL staining.
RESULTS
Hippo signaling was the top differentially methylated pathway in SLE versus control keratinocytes. SLE keratinocytes (n = 6) showed significant hypomethylation (Δβ = -0.153) and thus overexpression of the Hippo regulator WWC1 (P = 0.002). WWC1 overexpression increased LATS1/2 kinase activation, leading to YAP cytoplasmic retention and altered proapoptotic transcription in SLE keratinocytes. Accordingly, UVB-mediated apoptosis in keratinocytes could be enhanced by WWC1 overexpression or YAP-TEAD inhibition, mimicking SLE keratinocytes. Importantly, inhibition of LATS1/2 with either the chemical inhibitor TRULI or siRNA effectively eliminated enhanced UVB-apoptosis in SLE keratinocytes.
CONCLUSION
Our work unravels a novel driver of photosensitivity in SLE: overactive Hippo signaling in SLE keratinocytes restricts YAP transcriptional activity, leading to shifts that promote UVB apoptosis.
目的
光敏性是红斑狼疮（SLE）最常见的表现之一，但发病机制尚不清楚。SLE 患者正常表皮表现出紫外线 B（UVB）驱动的细胞死亡，这种细胞死亡在细胞培养中持续存在。在这里，我们研究了表观遗传修饰和 Hippo 信号在 SLE 角质形成细胞中增强的 UVB 诱导细胞凋亡中的作用。
方法
我们分析了来自 SLE 患者的培养角质形成细胞与来自健康对照者的角质形成细胞（每组 n=6）中的 DNA 甲基化。使用免疫印迹和免疫荧光法在培养的角质形成细胞中验证蛋白质表达。通过慢病毒载体生成过表达 WWC1 的永生化角质形成细胞系。使用大肿瘤抑制因子激酶 1/2（LATS1/2）抑制剂（TRULI）和小干扰 RNA（siRNA）抑制 WWC1 驱动的变化。通过过表达表达四环素诱导的绿色荧光蛋白标记蛋白的 N/TERT 细胞系抑制 Yes 相关蛋白（YAP）和转录增强子相关结构域（TEAD）之间的相互作用，该蛋白抑制 YAP-TEAD 结合（TEADi）。通过用 cleaved caspase 3/7 和 TUNEL 染色评估细胞凋亡。
结果
Hippo 信号通路是 SLE 与对照角质形成细胞相比差异甲基化的首要途径。SLE 角质形成细胞（n=6）表现出明显的低甲基化（Δβ=-0.153），因此 Hippo 调节剂 WWC1 过表达（P=0.002）。WWC1 过表达增加了 LATS1/2 激酶的激活，导致 SLE 角质形成细胞中 YAP 细胞质保留和改变的促凋亡转录。因此，WWC1 过表达或 YAP-TEAD 抑制可增强角质形成细胞中的 UVB 介导的凋亡，模拟 SLE 角质形成细胞。重要的是，用化学抑制剂 TRULI 或 siRNA 抑制 LATS1/2 可有效消除 SLE 角质形成细胞中增强的 UVB 凋亡。
结论
我们的工作揭示了 SLE 光敏性的新驱动因素：SLE 角质形成细胞中过度活跃的 Hippo 信号限制了 YAP 的转录活性，导致促进 UVB 凋亡的转变。
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