Regulation of transcription factor function by proteolysis.
Regulation of transcription factor function by proteolysis.
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通过蛋白水解调节转录因子功能。
DOI：
10.1042/bst0250498
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发表时间：
1997
影响因子：
3.9
通讯作者：
P. King
中科院分区：
文献类型：
--
作者：
S. Goodbourn;P. King
通过排除转录激活因子IRF-1[32,33]。然而，随后的基因破坏实验表明，IRF-2对于1-干扰素[34]的基础抑制似乎是完全没有必要的。尽管结果如此，IRF-2可能在病毒感染后期发生的转录关闭中发挥作用。IRF-2（表观分子质量为57 kDa）在病毒感染期间被切割，留下一个n端片段（IRF-2*分子质量为26 kDa），该片段仍然可以与DNA结合[35-381]（见图1）。这种截断的产物与全长IRF-2具有完全不同的性质。虽然IRF-2只能以短暂的方式与DNA结合，但IRF-2*形成稳定的复合物，其关闭率超过90 min[38]，并且在转染实验中，IRF-2"是比全长IRF-2[38]更强的转录抑制因子。由于强烈抑制的IRF-2”在1-干扰素mRNA合成接近峰值时开始形成，并在1-干扰素mRNA水平下降期间继续积累，因此IRF-2*可能是作为诱导后抑制因子发挥作用。该模型与irf -2- i敲除小鼠显示1-干扰素表达延迟关闭[34]的观察结果一致。为了绘制IRF-2的切割位点，我们将体外产生的截断IRF-2的电泳迁移率与体内的切割产物（IRF-2*）进行了比较，从而将切割位点定位在163和181[38]氨基酸之间。对该区域的氨基酸序列进行目视检查以寻找潜在的切割位点，发现氨基酸168和171之间的基序为Glu-Val-Ala-Asp-Ser，该序列与半胱氨酸蛋白酶ICE家族成员pcp -32的一致序列相似（见[39]）。阻断NF KB活化的蛋白酶抑制剂不抑制IRF-2的切割。然而，当细胞用ice样蛋白酶抑制剂z - val - asp -氟甲基酮处理时，IRF-2*的生成被完全阻断。这种治疗也导致了1-干扰素mRNA合成的延长和增强，这与IRF-2的作用一致。同样的处理不能阻断NFKB的诱导。为了证实ice样蛋白酶在IRF-2*生成中的作用，我们将170号氨基酸上的天冬氨酸残基改变为丙氨酸。这种改变形式的IRF-2在诱导过程中没有被切割。IRF-2的另一个功能可能是调节细胞生长。有趣的是，IRF-2在GI-S转化过程中被认为是组蛋白H4基因F0108转录的激活因子。由于全长IRF-2不能作为/)干扰素表达的激活因子[35,38]，这一结果表明，IRF-2可能根据启动子上下文作为抑制因子或激活因子。人工截断小鼠IRF-2表明，去除c端59个氨基酸揭示了一个强的转激活结构域[42]（见图1），我们已经用人类IRF-2证实了这一结果。因此，组蛋白H4基因FOl08的转录激活可能利用了IRF-2的这个结构域，尽管尚不清楚该结构域是如何被揭开的。一种可能是翻译后修饰；例如，c端可能在GI-S转变时发生修饰，从而允许激活域发挥作用，Vaughan等人报道了一种与IRF-2免疫相关的G -s特异性因子的存在。我们没能
500 tion by excluding the transcriptional activator IRF-1 [32, 33]. However, subsequent gene disruption experiments have shown that IRF-2 appears to be wholly dispensable for basal repression of 1-interferon [34]. Despite this result, IRF-2 may play a role in the turn-off of transcription that occurs late on during viral infection. IRF-2 (apparent molecular mass of 57 kDa) is cleaved during viral infection to leave an N-terminal fragment (IRF-2* of molecular mass 26 kDa) that can still bind to DNA [35-381 (see Figure 1). This truncated product has profoundly different properties from full-length IRF-2. Whereas IRF-2 can only bind to DNA in a transient manner, IRF-2* forms stable complexes that have off rates in excess of 90 min [38], and in transfection experiments IRF-2" is a much stronger repressor of transcription than full-length IRF-2 [38]. Since the strongly repressing IRF-2" begins to be formed at a time when 1-interferon mRNA synthesis is near its peak, and continues to accumulate during the decline in 1-interfercn mRNA levels, it is probable that IRF-2* functions as a post-induction repressor. This model is consistent with the observation that IRF-2-I-knock-out mice show delayed shut off of 1-interferon expression [34]. In an attempt to map the cleavage site of IRF-2, the electrophoretic mobility of truncated forms of IRF-2 generated in vitro were compared with the in vivo cleavage product (IRF-2*), allowing the cleavage site to be mapped to between amino acids 163 and 181 [38]. A visual inspection of the amino acid sequence in this region for potential cleavage sites revealed the motif Glu-Val-Ala-Asp-Ser between amino acids 168 and 171, a sequence that is similar to the consensus for CPP-32, a member of the ICE family of cysteine proteases (reviewed in [39]). Protease inhibitors that block activation of NF KB did not inhibit cleavage of IRF-2. However, when cells were treated with the ICE-like protease inhibitor Z-Val-Asp-fluoromethylketone, the generation of IRF-2* was completely blocked. This treatment also caused a prolonged and intensified synthesis of 1-interferon mRNA, consistent with the proposed role for IRF-2". The same treatment failed to block the induction of NFKB. To confirm the role of ICE-like proteases in the production of IRF-2*, we altered the aspartic acid residue at amino acid 170 to an alanine. This altered form of IRF-2 was not cleaved during induction. An additional function of IRF-2 may be to regulate cell growth [40]. Intriguingly, IRF-2 has been proposed to function as an activator of the histone H4 gene F0108 transcription during the GI-S transition [41]. Since full-length IRF-2 cannot act as an activator of/)-interferon expression [35, 38], this result suggests that IRF-2 may act as a repressor or activator depending on promoter context. Artificial truncation of mouse IRF-2 demonstrated that removal of the C-terminal 59 amino acids unveiled a strong transactivation domain [42](see Figure l), a result that we have confirmed using human IRF-2. Thus it is possible that transcriptional activation of the histone H4 gene FOl08 utilizes this domain of IRF-2, although it is not clear how the domain would become unmasked. One possibility is post-translational modification; for example, the C-terminus might become modified at the GI-S transition allowing the activation domain to function, and Vaughan et al.[41] have reported the existence of a G,-S-specific factor that is immunologically related to IRF-2. We have failed to