Structural basis for CEP192-mediated regulation of centrosomal AURKA

Abstract

Aurora kinase A (AURKA) performs critical functions in mitosis. Thus, the activity and subcellular localization of AURKA are tightly regulated and depend on diverse factors including interactions with the multiple binding cofactors. How these different cofactors regulate AURKA to elicit different levels of activity at distinct subcellular locations and times is poorly understood. Here, we identified a conserved region of CEP192, the major cofactor of AURKA, that mediates the interaction with AURKA. Quantitative binding studies were performed to map the interactions of a conserved helix (Helix-1) within CEP192. The crystal structure of Helix-1 bound to AURKA revealed a distinct binding site that is different from other cofactor proteins such as TPX2. Inhibiting the interaction between Helix-1 and AURKA in cells led to the mitotic defects, demonstrating the importance of the interaction. Collectively, we revealed a structural basis for the CEP192-mediated AURKA regulation at the centrosome, which is distinct from TPX2-mediated regulation on the spindle microtubule.

Centrioles generate two scaffolds with distinct biophysical properties to build mitotic centrosomes

“…This interaction comprised two independent interfaces involving two short regions of Spd-2 (Spd-2229-250 and Spd-2291-310), which we term AurA b inding d omain (ABD) 1 and 2, respectively, wrapping around the surface of the kinase domain. Interestingly, Spd-2-ABD1 and Spd-2-ABD2 are predicted to bind to similar regions on AurA/AURKA as human CEP192 (Park et al, 2023) and human TPX2 (Bayliss et al, 2003), respectively (Figure 5A; Figure S3).…”

“…Using the iPTM score as an initial assessment of each predicted interaction, we identified the interaction between the AurA kinase domain (AurA155-421) and Spd-2291-310 (subsequently named ABD2) as the strongest hit (iPTM=0.769). Shortly afterwards, a potential interaction interface between HsAURKA and HsCEP192 was identified (Park et al, 2023). This was different to the potential interaction we initially identified, but in a screen using ColabFold v1.5.5 in which AlphaFold2-multimer V3 was embedded where we used the N-terminal half of Spd-2 (Spd-21-Spd-2650) with the AurA kinase domain (AurA155-421), an interaction interface similar to the one identified in humans was found with Spd-2229-250 (later named ABD1).…”

“…In support of this possibility, halving the genetic dose of Spd-2 reduced TACC scaffold assembly in a similar manner to halving the dose of AurA (Figure 3). In vertebrates, the interaction between CEP192 and AURKA is well characterised (Joukov et al, 2010(Joukov et al, , 2014Meng et al, 2015) and the crystal structure of an interaction interface has recently been described (Park et al, 2023). It is unclear, however, if invertebrate versions of these proteins interact (Joukov and De Nicolo, 2018).…”