Linking cell-matrix adhesions and microtubules: the role of KANK proteins

Abstract

In this thesis, we characterized the KANK family proteins and their role as mediators of the crosstalk between the microtubule (MT) cytoskeleton and integrin-mediated cell-matrix adhesions. We tried to better understand how KANKs are recruited to FAs, how they interact with talin and what other factors contribute to their function and subcellular localization. In Chapter 2, we establish KANK family proteins as novel binding partners of the FA protein talin. Thereby KANKs form the link between FAs and the CMSC. Further, we could show that the talin-KANK interaction is required for proper formation of the CMSC at the cortex and that perturbation of this connection leads to MT overgrowth. In Chapter 3, we describe the interaction between talin and KANK1 in more detail. We found that the interaction between talin rod domain R7 and the KANK1 KN domain is force-regulated and that it can withstand forces in the piconewton range under certain force geometry. In Chapter 4, we focus on the subcellular localization of KANK1 and explain how the clustering of KANKs around FAs is mediated. We were able to demonstrate that this localization is not due to the coiled coil nor the ankyrin repeat domains but due to the linker region L2 of KANK1. In Chapter 5, we describe the differences between KANK1 and KANK2 with a particular focus on their different subcellular localizations and their role in cell migration. KANK1 is predominantly localized at the cell periphery whereas KANK2 is more concentrated in the central region. We were able to show that the localization of KANK2 is mediated by its KN domain and binding to talin. Further, have seen that depletion of both KANK1 and 2 does not affect cell migration of HeLa cells. On the other hand, loss of KANK2 significantly hampered cell migration of SUM159PT breast cancer cells. In Chapter 6, we discuss how KANK family proteins regulate the crosstalk between MTs and FAs. We combine our work on KANKs as linkers between FAs and CMSC (Chapter 2), the force-regulated interaction between talin and KANKs (Chapter 3), the role of KANK1’s linker region L2 in localizing KANK1 around FAs (Chapter 4), and the role of KANK family proteins in cell migration (Chapter 5) and put them into broader perspective. Further, we discuss some of the partially contradictory findings on KANK family proteins from us and others. We also present some preliminary data on KANK family proteins in endothelial cells and discuss an involvement of KANKs in ECM reorganization (fibronectin fibrillogenesis) and angiogenesis.