POST-TRANSLATIONAL MODIFICATIONS IN MAMMARY GLAND DEVELOPMENT AND MAMMARY TUMOR PROGRESSION

Breast cancer is one of the most common cancers in women. Estrogen receptor α (ERα) signaling and p53 signaling have important roles in breast cancer progression. Therefore, post-translational modifications of ERα and p53 play critical roles in breast cancer. The overall aim of this thesis is to characterize the role of RING-finger protein 31 (RNF31) on ERα and p53 signaling and the function of P21-activated kinase 4 (PAK4) on ERα signaling. Moreover, the role of PAK4 in mouse mammary development and mammary tumor progression was also analyzed. In the first study, RNF31 was shown to active and stabilize ERα, and subsequently to increase estrogen-stimulated cell proliferation in breast cancer cells. In breast cancer clinical databases, the gene expression of RNF31 and ERα target genes were correlated. The suggested mechanism is that RNF31 interacts ERα via the RBR domain and facilitate ERα mono-ubiquitination. In the second study, RNF31 depletion was shown to increase the gene expression of p53 target genes. RNF31 depletion caused cycle arrest and cisplatin-induced apoptosis in a p53dependent manner in breast cancer cells. Depletion of RNF31 increased p53 protein levels and the mRNA levels of its downstream target genes. The suggested mechanism is that RNF31 interacts with the p53/MDM2 complex and stabilizes MDM2 and consequently facilitates p53 poly-ubiquitination and degradation. In the third study, high PAK4 expression level was correlated with poor tamoxifen response in breast cancer patients in clinical databases, based on analysis of available mRNA expression. In MCF-7 cells, PAK4 overexpression promoted tamoxifen resistance, while PAK4 inhibition sensitized tamoxifen-resistant breast cancer cells to tamoxifen. Mechanistically, we identified a regulatory positive feedback loop, where PAK4 acts as a downstream target gene of ERα; while PAK4 can phosphorylate ERα at Ser305, thereby increasing ERα protein stability and activating ERα signaling. In conclusion, PAK4 may be a suitable target for tamoxifen resistance in breast cancer. In the fourth study, we elucidated the function of PAK4 in mammary development and mammary tumor progression in vivo. We observed no difference in mammary gland development between control mice and PAK4 conditional knockout mice. To test the role of PAK4 in mammary tumor development, conditional depletion of PAK4 was introduced in the MMTV-PyMT breast cancer mouse model. Importantly, conditional PAK4 depletion caused an increased tumor latency in MMTV-PyMT mice, indicating a role for PAK4 in early mammary tumor development. LIST OF SCIENTIFIC PAPERS I. J Zhu, C Zhao, A Kharman-Biz, T Zhuang, P Jonsson, N Liang, C Williams, C-Y Lin, Y Qiao, K Zendehdel, S Stromblad, E Treuter, K Dahlman-Wright. The atypical ubiquitin ligase RNF31 stabilizes estrogen receptor α and modulates estrogen-stimulated breast cancer cell proliferation. Oncogene 01/2014; DOI:10.1038/onc.2013. 573; 33(4340-4351). II. Jian Zhu, Chunyan Zhao, Ting Zhuang, Philip Jonsson, Cecilia Williams, Staffan Stromblad, Karin Dahlman-Wright. RING finger protein 31 (RNF31) promotes p53 degradation in breast cancer cells. Oncogene 07/2014; DOI: 10.1038/onc.2015. 260. III. Ting Zhuang*, Jian Zhu*, Zhilun Li, Julie Lorent, Karin Dahlman-Wright, Staffan Stromblad. Pharmacological targeting of p21-activated kinase-4 inhibits estrogen receptor alpha signaling and restores tamoxifen-sensitivity in breast cancer cells. (Manuscript) * Equal contributors IV. Ting Zhuang, Tania Costa, Parisa Rabieifar, Staffan Stromblad. Increased MMTV-PyMT mammary tumor latency by MMTV-Cre-driven conditional gene depletion of p21-activated kinase 4. (Manuscript) LIST OF ADDITIONAL PUBLICATIONS (NOT INCLUDED IN THE THESIS) I. Yichun Qiao, Chiou-Nan Shiue, Jian Zhu, Ting Zhuang, Philip Jonsson, Anthony P H Wright, Chunyan Zhao, Karin Dahlman-Wright: AP-1-mediated chromatin looping regulates ZEB2 transcription: new insights into TNFalphainduced epithelial-mesenchymal transition in triple-negative breast cancer. Oncotarget, 2015. 6(10): p. 7804-14. CONTENTS