No abstract

WW domain tandem-containing proteins such as KIBRA, YAP, and MAGI play critical roles in cell growth and polarity via binding to and positioning target proteins in specific subcellular regions. An immense disparity exists between promiscuity of WW domain-mediated target bindings and specific roles of WW domain proteins in cell growth regulation. Here, we discovered that WW domain tandems of KIBRA and MAGI, but not YAP, bind to specific target proteins with extremely high affinity and exquisite sequence specificity. Via systematic structural biology and biochemistry approaches, we decoded the target binding rules of WW domain tandems from cell growth regulatory proteins and uncovered a list of previously unknown WW tandem binding proteins including β-Dystroglycan, JCAD, and PTPN21. The WW tandem-mediated target recognition mechanisms elucidated here can guide functional studies of WW domain proteins in cell growth and polarity as well as in other cellular processes including neuronal synaptic signaling.

SUMMARY The canonical mammalian Hippo pathway contains a core kinase signaling cascade requiring upstream MST to form a stable complex with SAV1 in order to phosphorylate the downstream LATS/MOB complex. Though SAV1 dimerization is essential for the trans-activation of MST, the molecular mechanism underlying SAV1 dimerization is unclear. Here, we discover that the SAV1 WW tandem containing a short Pro-rich extension immediately following the WW tandem (termed as “WW12ex”) forms a highly stable homodimer. The crystal structure of SAV1 WW12ex reveals that the Pro-rich extension of one subunit binds to both WW domains from the other subunit. Thus, SAV1 WW12ex forms a domain-swapped dimer instead of a WW2 homodimerization-mediated dimer. The WW12ex-mediated dimerization of SAV1 is required for the MST/SAV1 complex assembly and MST kinase activation. Finally, we show that several cancer-related SAV1 variants disrupt SAV1 dimer formation, and thus, these mutations may impair the tumor-suppression activity of SAV1.