Structural Basis for Rab8a GTPase Recruitment of RILPL2 via LRRK2-Mediated Phosphorylation of Switch 2

Rab8a GTPase is associated with the dynamic regulation of membrane protrusions in polarized cells.  Rab8a is one of several Rab-family GTPases that are substrates of leucine-rich repeat kinase 2 (LRRK2), a serine/threonine kinase that is linked to inherited Parkinson's disease.  Rab8a is phosphorylated at T72 (pT72) in its switch 2 helix and the post-translational modification facilitates phospho-Rab8a (pRab8a) interactions with RILPL2, which subsequently regulates ciliogenesis. Here we report the crystal structure of pRab8a in complex with the phospho-Rab binding domain of RILPL2.  The complex is a heterotetramer with RILPL2 forming a central alpha-helical dimer that bridges two pRab8a molecules.  The N-termini of the a-helices cross over to form an X-shaped cap (X-cap) that enables electrostatic interactions between Arg residues from RILPL2 and the phosphate moiety from pT72.  RILPL2 residues in the X-cap that are critical for pRab8a binding are conserved in the RILP family of effector proteins.  These structural insights revealed that JIP3 and JIP4 also possess a phospho-Rab binding domain similar to RILPL2 including the conserved residues that bind to pRab8A. This prompted us to analyze these proteins and we found that JIP3 and JIP4 interact specifically with LRRK2-phosphorylated Rab10, suggesting a general mode of recognition for phosphorylated Rab GTPases by phospho-specific effectors.