Kim Eun-Joo, Park Jong-Sup, Um Soo-Jong
Department of Molecular Biology, Dankook University, Gyeonggi-do 448-701, Republic of Korea.
Biochem Biophys Res Commun. 2008 Dec 19;377(3):952-6. doi: 10.1016/j.bbrc.2008.10.092. Epub 2008 Oct 26.
To characterize the regulatory mechanism of the interferon regulatory factor (IRF) family, we performed yeast two-hybrid screening with IRF-2 and isolated the small ubiquitin-related modifier (SUMO)-conjugating enzyme Ubc9, which also interacts with other IRF family members IRF-1 and ICSBP. Subsequent assays indicated that among the IRF family members, only IRF-1 interacts with SUMO-1 through its transcriptional activation domain. The interaction between IRF-1 and SUMO-1 was confirmed in vitro by GST pull-down assays and in vivo by co-localization assays. Furthermore, this interaction led to the Ubc9-mediated sumoylation of IRF-1 in vitro and in vivo. Transient transfection assays revealed that Ubc9 or SUMO inhibits the transcriptional activity of IRF-1 in a dose-dependent manner. Finally, Ubc9 and SUMO cooperate in the transcriptional repression of IRF-1. Taken together, these observations suggest that Ubc9 functions as a transcriptional repressor of IRF-1 by inducing sumoylation, and that this effect may be required for the physiological activity of IRF-1.
为了阐明干扰素调节因子（IRF）家族的调控机制，我们利用IRF-2进行酵母双杂交筛选，并分离出小泛素相关修饰物（SUMO）结合酶Ubc9，它也与其他IRF家族成员IRF-1和ICSBP相互作用。随后的实验表明，在IRF家族成员中，只有IRF-1通过其转录激活结构域与SUMO-1相互作用。通过GST下拉实验在体外和共定位实验在体内证实了IRF-1与SUMO-1之间的相互作用。此外，这种相互作用导致了体外和体内Ubc9介导的IRF-1的SUMO化。瞬时转染实验表明，Ubc9或SUMO以剂量依赖的方式抑制IRF-1的转录活性。最后，Ubc9和SUMO在IRF-1的转录抑制中协同作用。综上所述，这些观察结果表明，Ubc9通过诱导SUMO化作为IRF-1的转录抑制因子发挥作用，并且这种作用可能是IRF-1生理活性所必需的。
