Function of MM-1, a novel c-Myc-binding protein, as a tumor suppressor

1).一条新的c-Myc转录抑制途径及其靶基因的鉴定我们发现MM-1与转录辅阻遏子Tif1β结合，并且MM-1向c-Myc招募了包括HDAC1和mSin3在内的辅阻遏子复合体，从而抑制了c-Myc的转录活性。为了确定这一途径的靶基因，首先建立了一个含有显性-阴性形式的TIF1β的MycER细胞系，其中辅阻遏子复合体不被招募到c-Myc。然后应用DNA微阵列方法鉴定与MycER细胞相比在该系中表达上调的基因。利用该系统，我们鉴定c-fms癌基因为c-Myc-MM-1抑制基因。此外，我们还发现Wint-4基因是MM-1的转录抑制靶基因。在MM-1-基因敲除细胞中，发现Wint信号被激活，导致c-myc表达被激活。2)c-Myc新的降解途径的鉴定我们还鉴定了Elongin B、26S蛋白酶体亚单位Rpt3和Rpn12为MM-1相关蛋白。由于这些蛋白质具有泛素化和蛋白质降解的功能，我们随后测试了MM-1在c-Myc降解中发挥作用的可能性。结果表明，MM-1通过新型E3泛素连接酶复合体Spk2、cullin 1、Elongon B促进c-Myc的降解，并利用针对每个基因的siRNA证实了这种现象在体内发生。这些结果表明MM-1是负调控c-Myc功能的关键蛋白，突变使MM-1的这些功能丧失导致c-Myc形成肿瘤。

1).Identification of a novel transrepression pathway of c-Myc and its target geneWe have found that MM-1 bound to TIF1β, a transcriptional corepressor, and that MM-1 recruited a corepressor complex, including HDAC1 and mSin3, to c-Myc, thereby leading to repressing c-Myc transcription activity. To identify target genes to this pathway, a MycER cell line harboring a dominant-negative form of TIF1β, in which the corepressor complex was not recruited to c-Myc, was first established. DNA-microarray method was then applied to identify genes whose expressions were upregulated in this line compared to those in MycER cells. By this system we identified c-fms oncogene as the c-Myc-MM-1 repressed gene. Furthermore, we also identified the Wint-4 gene as a transcriptional repression target of MM-1. In MM-1-knockdown cells, the Wint signal was found to be activated, resulting in activation of c-myc expression.2).Identification of a novel degradation pathway of c-MycWe have also identified Elongin B, 26Sproteasome subunits Rpt3 and Rpn12 as MM-1-associated proteins. Since these proteins functions for ubiquitination and degradation of proteins, we then tested a possibility that MM-1 plays a role in c-Myc degradation. The results indicate that MM-1 stimulated c-Myc degradation through the novel E3 ubiquitin ligase complex, including Spk2, Cullin 1, Elongon B, and these phenomena were confirmed to occur in vivo using siRNAs targeting each genes.These findings indicate that MM-1 is a key protein that negatively regulates c-Myc function and that lost of these functions of MM-1 by mutations leads to tumor formation by c-Myc.