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

1).一条新的c-Myc反式阻遏途径及其靶基因的鉴定我们发现MM-1与转录辅阻遏物TIF 1 β结合，并且MM-1向c-Myc募集辅阻遏物复合物，包括HDAC 1和mSin 3，从而导致c-Myc转录活性被抑制。为了鉴定该途径的靶基因，首先建立了一种携带显性阴性形式的TIF 1 β的MycER细胞系，其中辅阻遏物复合物不被募集到c-Myc。然后应用DNA微阵列方法来鉴定在该细胞系中与MycER细胞中相比表达上调的基因。通过该系统我们鉴定出c-fms癌基因为c-Myc-MM-1抑制基因。此外，我们还确定了Wint-4基因作为MM-1的转录抑制靶标。在MM-1敲低的细胞中，发现Wint信号被激活，导致c-myc表达的激活。2）c-myc的新降解途径的鉴定我们还鉴定了延伸蛋白B、26 S蛋白酶体亚基Rpt 3和Rpn 12作为MM-1相关蛋白。由于这些蛋白质的功能是蛋白质的泛素化和降解，因此我们测试了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泛素连接酶复合物（包括Spk 2、Cullin 1、Elongon B）刺激c-Myc降解，并且使用靶向每个基因的siRNA证实这些现象在体内发生。这些发现表明MM-1是负调节c-Myc功能的关键蛋白，并且通过突变导致MM-1的这些功能的丧失导致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.