Study On The Roles And Mechanism Of HPD In The Tumorigenesis And Development Of Lung Cancer

This thesis is divided into two parts. One part is about the roles of HPD in the tumorigenesis and development of lung cancer and the mechanism of HPD on regulating the malignant phenotype of lung cancer. The other part is about the mechanism of THAP11 on regulating hematopoietic differentiation.In the first part, we focuses on the protein named HPD, which is one of the key enzymes involved in tyrosine metabolism. Previous study suggested that HPD was expressed sepcifically in liver and kidney. In the present study, we investigated the expression level of HPD in lung caner patients using tissue microarray and our results showed that the expression level of HPD in carcinoma tissues of lung cancer patients was significantly higher than adjacent tissues, and the up-regulation of HPD was confirmed by ELISA. To examine the roles of HPD in lung cancer, we established some lung cancer cell lines stably overexpressing HPD or HPD RNAi and found that overexpression of HPD didn’t affect the growth and proliferation of lung cancer cells but HPD knockdown inhibited the survival of lung cancer cells and reduced the tumorigenicity in vivo,indicating that HPD is one key factor involved in the growth and survival of lung cancer. Furthermore, overexpression of HPD enhanced the migration activity of several lung cancer cells while HPD knockdown inhibited the migration activity of lung cancer cells and reduced the metastasis in nude mice,which suggested that HPD promotes the invasion and metastasis of lung cancer cells. We also examined the effects of HPD on the sensitivity of lung cancer cells to drugs for treating lung cancer and our results showed that overexpression of HPD reduced the sensitivity of lung cancer cells to CDDP, Etopside and Gefitinib,and kncokdown of HPD enhances the sensitivity of A549 to drugs such as CDDP and Gefitinib with increased apoptosis. In addition, we also investigated the mechanism of HPD on regulating the phenotypes of lung cancer cells. We found that HPD enzyme inhibitor Nitisinone did not affect the proliferation and migration of lung cancer cells, suggesting that the effects of HPD on lung cancer cells didn’t depend on its enzyme activity. Based on the previous study, we also found that HPD activated NF-кB signaling pathway with promoting the degradation of IкBα and the nucleus translocation of p65, and regulating the downstream target genes of NF-кB, such as c-Myc, MMP9 and CyclinD1.These data suggest that HPD regulates human lung cancer survival, apoptosis and migration and positively regulates NF-?B signal pathway.In the second part, we focus on the transcription factor THAP11, which has been reported to play key roles in cell growth, embryogenesis, and ES cell pluripotency. Our previous results showed that THAP11 is a noval transcription factor involved in regulating erythroid and megakaryocytic differentiation of hematopoietic cells, but the mechanism is not clear. We investigated the mechanism of THAP11 on hematopoietic differentiation in this part. The results showed that during hemin-induced erythroid differentiation of K562 cells, THAP11 overexpression led to the down-regulation of erythroid transcription factor c-Myc and up-regulation of megakaryocytic transcription factor Fli 1. Bioinformatic analysis suggested that there are several potential THAP11 binding sites on the promoters of these genes. ChIP analysis suggested that THAP11 directly binded to specific sites on the promoter of various hematopoietic transcription factor including GATA-2, c-Myb, Fli 1 and c-Myc. These results indicated that THAP11 might regulate the expression levels of several key hematopoietic transcription factors directly. We further investigated the effects of THAP11 on histone deacetylation in K562 cells during megakaryocytic differentiation. The data suggested that the acetylation levels of H3K9 and H3K18 were not affected by THAP11 overexpression. Interestingly, we also provided evidence that THAP11 interacts with GATA-2, suggesting a potential regulation of THAP11 on GATA-2 activity. These findings suggest that THAP11 regulated erythroid/megakaryocytic differentiation probably by modifying some key erythroid or megakaryocytic genes.