Sublytic C5b-9 induces TIMP3 expression by glomerular mesangial cells via TRAF6-dependent KLF5 K63-linked ubiquitination in rat Thy-1 nephritis
Sublytic C5b-9 induces TIMP3 expression by glomerular mesangial cells via TRAF6-dependent KLF5 K63-linked ubiquitination in rat Thy-1 nephritis
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DOI：
10.1016/j.intimp.2023.110970
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发表时间：
2023-09-23
影响因子：
5.6
通讯作者：
Wang，Yingwei
中科院分区：
文献类型：
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作者：
Ying，Shuai;Liu，Longfei;Wang，Yingwei
大鼠Thy-1肾炎（Thy-1N）是研究人系血管增殖性肾小球肾炎（MsPGN）的实验模型，其病理特征为肾小球系膜细胞（GMC）增殖和细胞外基质（ECM）积累。虽然我们已经证实Thy-1N大鼠的肾脏病变依赖于亚溶C5b-9， ECM积累与基质金属蛋白酶（TIMP）组织抑制剂抑制基质金属蛋白酶（MMP）活性有关，但亚溶C5b-9是否能诱导Thy-1N大鼠的GMC产生TIMP及其机制尚不清楚。在本研究中，我们证明了TIMP3、krϋppel-like转录因子5 （KLF5）和肿瘤坏死因子受体相关因子6 （TRAF6）的表达在Thy-1N大鼠肾组织（体内）和暴露于亚溶C5b-9的GMC（体外）中同时上调。进一步的机制探索发现，KLF5和TRAF6作为两个上游分子，可以通过结合TIMP3启动子的−1801nt ~−1554nt （GGGGAGGGGC）和−228nt ~−46nt （GCCCCGCCCC）同一区域，诱导TIMP3基因转录。在此过程中，TRAF6介导KLF5在K99和K100位点k63连锁泛素化，增强KLF5的核定位和与TIMP3启动子的结合，增强其基因活化。此外，体内实验表明，沉默KLF5、TRAF6或TIMP3基因可显著减轻Thy-1N大鼠肾脏KLF5 k63连锁泛素化或TIMP3诱导、ECM积累等病理改变。此外，上述这些蛋白在MsPGN患者肾组织中的阳性表达与ECM积累及其相关性也得到了证实。总之，我们的研究结果表明KLF5和TRAF6在亚裂解c5b -9触发的TIMP3基因的转录和表达中发挥促进作用，这可能为大鼠Thy-1N和人MsPGN的机制提供新的见解。
Rat Thy-1 nephritis (Thy-1N) is an experimental model for studying human mesangioproliferative glomerulonephritis (MsPGN), and its pathological features are glomerular mesangial cell (GMC) proliferation and extracellular matrix (ECM) accumulation. Although we have confirmed that renal lesions of Thy-1N rats are sublytic C5b-9-dependent, and ECM accumulation is related to tissue inhibitor of matrix metalloproteinase (TIMP) inhibiting matrix metalloproteinase (MMP) activity, whether sublytic C5b-9 can induce TIMP production by GMC in Thy-1N rat and the underlying mechanism remains unclear. In the study, we proved that the expressions of TIMP3, krϋppel-like transcription factor 5 (KLF5) and tumor necrosis factor receptor-associated factor 6 (TRAF6) were simultaneously up-regulated both in the renal tissues of Thy-1N rats (in vivo) and in the GMC exposed to sublytic C5b-9 (in vitro). Further mechanism exploration discovered that KLF5 and TRAF6 as two upstream molecules could induce TIMP3 gene transcription through binding to the same region i.e., −1801nt to −1554nt (GGGGAGGGGC) and −228nt to −46nt (GCCCCGCCCC) of TIMP3 promoter. In the process, TRAF6 mediated KLF5 K63-linked ubiquitination at K99 and K100 enhancing KLF5 nuclear localization and binding to TIMP3 promoter, augmenting its gene activation. Furthermore, the experiments in vivo exhibited that silencing KLF5, TRAF6 or TIMP3 gene could markedly lessen renal KLF5 K63-linked ubiquitination or TIMP3 induction, ECM accumulation and other pathological changes of Thy-1N rats. Besides, the positive expressions of above-mentioned these proteins and ECM accumulation and their correlation in the renal tissues of MsPGN patients were also demonstrated. Overall, our findings implicate that KLF5 and TRAF6 play a promoting role in sublytic C5b-9-triggered TIMP3 gene transcription and expression, which might provide a novel mechanistic insight into rat Thy-1N and human MsPGN.