Protein phosphatase 2a (PP2A) binds within the oligomerization domain of striatin and regulates the phosphorylation and activation of the mammalian Ste20-Like kinase Mst3

Abstract

BackgroundStriatin, a putative protein phosphatase 2A (PP2A) B-type regulatory subunit, is a multi-domain scaffolding protein that has recently been linked to several diseases including cerebral cavernous malformation (CCM), which causes symptoms ranging from headaches to stroke. Striatin association with the PP2A A/C (structural subunit/catalytic subunit) heterodimer alters PP2A substrate specificity, but targets and roles of striatin-associated PP2A are not known. In addition to binding the PP2A A/C heterodimer to form a PP2A holoenzyme, striatin associates with cerebral cavernous malformation 3 (CCM3) protein, the mammalian Mps one binder (MOB) homolog, Mob3/phocein, the mammalian sterile 20-like (Mst) kinases, Mst3, Mst4 and STK25, and several other proteins to form a large signaling complex. Little is known about the molecular architecture of the striatin complex and the regulation of these sterile 20-like kinases.ResultsTo help define the molecular organization of striatin complexes and to determine whether Mst3 might be negatively regulated by striatin-associated PP2A, a structure-function analysis of striatin was performed. Two distinct regions of striatin are capable of stably binding directly or indirectly to Mob3--one N-terminal, including the coiled-coil domain, and another more C-terminal, including the WD-repeat domain. In addition, striatin residues 191-344 contain determinants necessary for efficient association of Mst3, Mst4, and CCM3. PP2A associates with the coiled-coil domain of striatin, but unlike Mob3 and Mst3, its binding appears to require striatin oligomerization. Deletion of the caveolin-binding domain on striatin abolishes striatin family oligomerization and PP2A binding. Point mutations in striatin that disrupt PP2A association cause hyperphosphorylation and activation of striatin-associated Mst3.ConclusionsStriatin orchestrates the regulation of Mst3 by PP2A. It binds Mst3 likely as a dimer with CCM3 via residues lying between striatin's calmodulin-binding and WD-domains and recruits the PP2A A/C heterodimer to its coiled-coil/oligomerization domain. Residues outside the previously reported coiled-coil domain of striatin are necessary for its oligomerization. Striatin-associated PP2A is critical for Mst3 dephosphorylation and inactivation. Upon inhibition of PP2A, Mst3 activation appears to involve autophosphorylation of multiple activation loop phosphorylation sites. Mob3 can associate with striatin sequences C-terminal to the Mst3 binding site but also with sequences proximal to striatin-associated PP2A, consistent with a possible role for Mob 3 in the regulation of Mst3 by PP2A.

Misshapen-like kinase 1 (MINK1) Is a Novel Component of Striatin-interacting Phosphatase and Kinase (STRIPAK) and Is Required for the Completion of Cytokinesis

“…STRN4, a regulatory subunit of PP2A, directly associated with MINK1, and depletion of STRN4-induced multinucleated cells and inhibited abscission. Striatins-associated PP2A has been reported to dephosphorylate and inactivate serine/threonine kinase 24 (STK24), a member of the germinal center kinases that is known to complex with striatins (31). Similarly, we found that STRN4 could reduce the catalytic activity of MINK1 in the presence of structural and catalytic subunits of PP2A.…”

Striatins Contain a Noncanonical Coiled Coil That Binds Protein Phosphatase 2A A Subunit to Form a 2:2 Heterotetrameric Core of Striatin-interacting Phosphatase and Kinase (STRIPAK) Complex

“…Striatins are a novel family of Bٞ type regulatory subunits that directly interact with PP2A A subunits, and indirectly interact with some GCKs using adaptor molecules (Fig. 1A) (7)(8)(9)(10)(11)(12). Because inhibition of PP2A using okadaic acid causes phosphorylation of MOB and hyperphosphorylation of striatins, PP2A may thus regulate striatins and the associated MOB proteins to modify the cytoskeleton and its interactions with membrane structures (10,11).…”

RHO binding to FAM65A regulates Golgi reorientation during cell migration

“…GCKIII kinases, along with their interacting protein CCM3, are known to be constituents of a conserved multi-protein complex known as the striatin-interacting phosphatase and kinase (STRIPAK) complex (Goudreault et al, 2009). In addition to GCKIII kinases and CCM3, STRIPAK contains the catalytic and scaffolding subunits of protein phosphatase-2A (PP2A) along with a specific family of regulatory subunits known as striatins (STRN1, STRN3 and STRN4), as well as STR-interacting proteins STRIP1 Fig.…”

“…and STRIP2, Mob-domain-containing protein MOB3 and the membrane-anchoring protein SLMAP (Goudreault et al, 2009;Kean et al, 2011). The STRIPAK complex has been shown to negatively regulate MST3 and MST4 kinase activities, most likely by dephosphorylating their activation segments through PP2A (Madsen et al, 2015).…”