Role of PAK4 in pancreas development and breast cancer

Sammanfattning: PAK4 is a Rho GTPase-regulated serine/threonine kinase that possesses critical functions in embryonic, neuronal and vascular development, immune defense and cancer. Constitutive PAK4 depletion causes embryonic lethality and mice with conditional PAK4 gene depletion in the heart and in the central nervous system displayed serious organ defects. PAK4 is overexpressed and genetically amplified in cancer cell lines as well as in cancer patients, including pancreas and breast. PAK4 also regulates many cellular functions related to cancer progression, including cell proliferation, survival and cell cycle as well as cell morphology, adhesion and migration, which are dependent on the actin cytoskeleton. Therefore, better understanding of the role of PAK4 in development, cancer progression and the mechanism behind that could help us to find suitable strategies to treat cancer. In paper I, we elucidated the role of PAK4 in pancreas development. PAK4 knockout mice were born at Mendelian ratios. Morphological and immunohistochemical examinations and quantifications indicated that exocrine, endocrine and ductal compartments retained the normal proportions and distributions upon PAK4 gene depletion. In addition, body weight records and a glucose tolerance test revealed no differences between wild type and PAK4 knockout mice. Together, this suggests that PAK4 is dispensable for mouse pancreas development. This will facilitate future use of our Pdx1-Cre-driven conditional PAK4 knock out mouse model for testing in vivo potential functions of PAK4 in pancreatic disease models such as for pancreatitis and different pancreatic cancer forms. In paper II, we comprehensively characterized the human PAK4 interactome. We found that the PAK4 interactome was enriched in diverse protein networks, including the 14-3-3, proteasome, replication fork, CCT and Arp2/3 complexes. Moreover, we found that PAK4 interacts with and phosphorylates VCA domain at Ser484/Ser485 and promotes Arp2/3- dependent actin polymerization in vitro. Also, PAK4 ablation in vivo reduced N-WASP Ser484/Ser485 phosphorylation and altered the cellular balance between G- and F-actin as well as the actin organization. By presenting the PAK4 interactome, we here provide a powerful resource for further investigations and we also indicate a novel mechanism by which PAK4 may regulate actin cytoskeleton remodeling. In paper III, we established PAK4 as a protumourigenic regulator of breast cancer acting through the non-canonical NF-κB subunit RelB. Our results demonstrated that PAK4 was overexpressed in human breast cancer and associated with poor prognosis. We found that PAK4 inhibition arrested cancer cell growth by inducing several senescence-like features. RNA sequencing and subsequent mechanistic exploration revealed that PAK4 inhibited NF- κB signaling. Further, we identified RelB, a subunit of the non-canonical NF-κB family, as necessary for senescence-like growth arrest upon PAK4 depletion. Importantly, we pinpointed RelB as a direct substrate of PAK4 and mapped a PAK4 phosphorylation residue (S151) within the Rel-homology domain that directly influences RelB-DNA interactions and target gene expression. Taken together, these studies highlight the importance of PAK4 in cancer and provide new mechanisms and new views to understanding the role of PAK4 in cancer cell progression