A versatile Scaffold: The binding specificities of the Par3 PDZ domains mediate multiple interactions with polarity proteins

Abstract

The asymmetric distribution of RNA, lipids and proteins is the basis of cell polarity. Polarized cells are vital for the organization of multicellular organism. Malfunctions in the processes generating cell polarity are linked with cancer and developmental defects. For cell polarization, the PAR complex, consisting of atypical protein kinase C, Par3 and Par6, is essential. Par3 is the central scaffold of the PAR complex. Par3 comprises of an N-terminal oligomerization domain, three Postsynaptic density protein-95, Disk large, Zonula occludens 1 (PDZ) domains, a kinase binding domain and an unstructured C-terminus. Its PDZ domains are the major protein-protein interaction domains. However, a detailed analysis of their specificities towards PDZ binding motifs (PBMs) occurring in Par3 interaction partners in the environment of cell polarity is missing. Here, I present the structural basis of the interaction of Par3 with Par6. I identified a PBM in Par6 that is essential for Par3 interaction and interacts with the PDZ1 and PDZ3, but not the PDZ2 domains in vitro. Together with my coauthors, I showed that the Par6 PBM interacts with Par3 via a canonical PDZ:PBM interaction and functions together with the Par6 PDZ domain in Par6 localization in vivo. In addition, I investigated the specificities of the individual Par3 PDZ domains for cell polarity proteins. My analysis revealed a unique binding profile for the dmPar3 PDZ1 and PDZ2 domains, while the binding profile of the dmPar3 PDZ3 domain is very promiscuous and overlaps with the specificities of the other two Par3 PDZ domains. These overlapping specificities enable Par3 to mediate multivalent interactions and thereby enable Par3 to form large protein networks with many different cell polarity proteins. In a third project, I discovered a hitherto unknown short motif N-terminal of the third PDZ domain of dmPar3, denoted FID-motif. I was able to show that the FID-motif folds back onto the dmPar3 PDZ3 domain in close vicinity of the PBM binding groove thereby reducing the affinities of the PDZ3 domain towards various PBMs in polarity proteins. These reductions in affinity prevent a subset of the previous identified PDZ3 ligands to interact with the PDZ3 domain. Hence, the FID-motif seems to fine-tune the recruitment of PBM-carrying polarity proteins via the dmPar3 PDZ3 domain. The detailed analyses presented in this thesis provide important insights into the individual roles of the Par3 PDZ domains in the assembly of polarity protein complexes. I present new clues in regard of functional redundancies within the Par3 PDZ module and provide the further evidence for Par3 acting as a central scaffold of polarity protein networks. Therefore, the function of the Par3 protein during establishment, maintenance and disruption of cell polarity during development and the related process of cancer metastasis can be understood in greater detail.