Tyrosine phosphorylation of the E3 ubiquitin ligase TRIM21 positively regulates interaction with IRF3 and hence TRIM21 activity.
Tyrosine phosphorylation of the E3 ubiquitin ligase TRIM21 positively regulates interaction with IRF3 and hence TRIM21 activity.
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DOI：
10.1371/journal.pone.0034041
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发表时间：
2012
期刊：
影响因子：
3.7
通讯作者：
Jefferies CA
中科院分区：
文献类型：
--
作者：
Stacey KB;Breen E;Jefferies CA
患有系统性红斑狼疮（SLE）的患者具有升高的I型干扰素（IFN）水平，其与疾病活动性和严重性相关。TRIM 21是一种与SLE相关的自身抗原，已被鉴定为靶向转录因子IRF 3的泛素E3连接酶，以便在通过Toll样受体（TLR）检测到病毒和细菌感染后关闭和限制I型IFN的产生。然而，TRIM 21的活性如何在TLR下游调节尚不清楚。在这项研究中，我们证明TRIM 21是酪氨酸磷酸化后TLR 3和TLR 4的刺激，表明其活性可能是由酪氨酸磷酸化调节。使用Netphos，我们已经确定了三个被强烈预测为磷酸化的关键酪氨酸，其中两个在TRIM 21的人类和小鼠形式之间是保守的，在残基343、388和393处，所有这些都已经从酪氨酸突变为苯丙氨酸（Y343 F、Y388 F和Y393 F）。我们已经观察到TRIM 21的酪氨酸磷酸化仅发生在底物结合PRY/SPRY结构域中，并且TRIM 21作为IFN-β启动子活性的负调节剂起作用需要Y393，并且在较小程度上需要Y388。进一步的研究表明，将Y393突变为苯丙氨酸会抑制TRIM 21与其底物IRF 3相互作用的能力，从而为Y393在IFN-β启动子上缺乏活性提供了分子解释。我们的数据表明酪氨酸磷酸化在调节TLR 3和TLR 4下游TRIM 21活性中的新作用。鉴于TRIM 21在系统性自身免疫中的致病作用，这些发现对新型疗法的开发具有重要意义。
Patients suffering from Systemic Lupus Erythematous (SLE) have elevated type I interferon (IFN) levels which correlate with disease activity and severity. TRIM21, an autoantigen associated with SLE, has been identified as an ubiquitin E3 ligase that targets the transcription factor IRF3 in order to turn off and limit type I IFN production following detection of viral and bacterial infection by Toll Like Receptors (TLRs). However, how the activity of TRIM21 is regulated downstream of TLRs is unknown. In this study we demonstrate that TRIM21 is tyrosine phosphorylated following TLR3 and TLR4 stimulation, suggesting that its activity is potentially regulated by tyrosine phosphorylation. Using Netphos, we have identified three key tyrosines that are strongly predicted to be phosphorylated, two of which are conserved between the human and murine forms of TRIM21, at residues 343, 388, and 393, all of which have been mutated from tyrosine to phenylalanine (Y343F, Y388F, and Y393F). We have observed that tyrosine phosphorylation of TRIM21 only occurs in the substrate binding PRY/SPRY domain, and that Y393, and to a lesser extent, Y388 are required for TRIM21 to function as a negative regulator of IFN-β promoter activity. Further studies revealed that mutating Y393 to phenylalanine inhibits the ability of TRIM21 to interact with its substrate, IRF3, thus providing a molecular explanation for the lack of activity of Y393 on the IFN-β promoter. Our data demonstrates a novel role for tyrosine phosphorylation in regulating the activity of TRIM21 downstream of TLR3 and TLR4. Given the pathogenic role of TRIM21 in systemic autoimmunity, these findings have important implications for the development of novel therapeutics.
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