Striatin-interacting Phosphatase Research Articles

A Hippo Pathway-Related GCK Controls Both Sexual and Vegetative Developmental Processes in the Fungus Sordaria macrospora.

The supramolecular striatin-interacting phosphatases and kinases (STRIPAK) complex is conserved from yeast to human, and regulates a variety of key biological processes. In animals, this complex consists of the scaffold protein striatin, the protein phosphatase 2A, and kinases, such as germinal center kinase (GCK) III and GCKIV family members, as well as other associated proteins. The STRIPAK complex was identified as a negative regulator of the Hippo pathway, a large eukaryotic signaling network with a core composed of a GCK and a nuclear Dbf2-related kinase. The signaling architecture of the Hippo core resembles the fungal septation initiation network (SIN) that regulates cytokinesis in fission yeast as well as septation in filamentous fungi. In the filamentous model fungus Sordaria macrospora, core components of the STRIPAK complex have been functionally described and the striatin homolog PRO11 has been shown to interact with the GCK SmKIN3. However, the exact role of SmKIN3 in fungal development has not yet been fully elucidated. Here, we provide comprehensive genetic and functional analysis of SmKIN3 from S. macrospora. Using deletion mutants and site-directed mutagenesis, along with phenotypic and phylogenetic analysis, we provide compelling evidence that SmKIN3 is involved in fruiting body formation, hyphal fusion, and septation. Strains carrying the ATP-binding mutant SmKIN3K39R, as well as a double-deletion strain lacking SmKIN3 and the core STRIPAK subunit PRO11, also revealed severe developmental defects. Collectively, this study suggests that SmKIN3 links both the SIN and STRIPAK complex, thereby regulating multiple key cellular processes.

Striatin-1 is a B subunit of protein phosphatase PP2A that regulates dendritic arborization and spine development in striatal neurons

Striatin-1, a subunit of the serine/threonine phosphatase PP2A, is preferentially expressed in neurons in the striatum. As a member of the striatin family of B subunits, striatin-1 is a core component together with PP2A of a multiprotein complex called STRIPAK, the striatin-interacting phosphatase and kinase complex. Little is known about the function of striatin-1 or the STRIPAK complex in the mammalian striatum. Here, we identify a selective role for striatin-1 in striatal neuron maturation. Using a small hairpin RNA (shRNA) knockdown approach in primary striatal neuronal cultures, we determined that reduced expression of striatin-1 results in increased dendritic complexity and an increased density of dendritic spines, classified as stubby spines. The dendritic phenotype was rescued by co-expression of a striatin-1 mutant construct insensitive to the knockdown shRNA but was not rescued by co-expression of PP2A- or Mob3-binding deficient striatin-1 constructs. Reduction of striatin-1 did not result in deficits in neuronal connectivity in this knockdown model, as we observed no abnormalities in synapse formation or in spontaneous excitatory postsynaptic currents. Thus, this study suggests that striatin-1 is a regulator of neuronal development in striatal neurons.

STRIP1, a core component of STRIPAK complexes, is essential for normal mesoderm migration in the mouse embryo

Regulated mesoderm migration is necessary for the proper morphogenesis and organ formation during embryonic development. Cell migration and its dependence on the cytoskeleton and signaling machines have been studied extensively in cultured cells; in contrast, remarkably little is known about the mechanisms that regulate mesoderm cell migration in vivo. Here, we report the identification and characterization of a mouse mutation in striatin-interacting protein 1 (Strip1) that disrupts migration of the mesoderm after the gastrulation epithelial-to-mesenchymal transition (EMT). STRIP1 is a core component of the biochemically defined mammalian striatin-interacting phosphatases and kinase (STRIPAK) complexes that appear to act through regulation of protein phosphatase 2A (PP2A), but their functions in mammals in vivo have not been examined. Strip1-null mutants arrest development at midgestation with profound disruptions in the organization of the mesoderm and its derivatives, including a complete failure of the anterior extension of axial mesoderm. Analysis of cultured mesoderm explants and mouse embryonic fibroblasts from null mutants shows that the mesoderm migration defect is correlated with decreased cell spreading, abnormal focal adhesions, changes in the organization of the actin cytoskeleton, and decreased velocity of cell migration. The results show that STRIPAK complexes are essential for cell migration and tissue morphogenesis in vivo.

Fsr1, a striatin homologue, forms an endomembrane-associated complex that regulates virulence in the maize pathogen Fusarium verticillioides.

Fsr1, a homologue of mammalian striatin, containing multiple protein-binding domains and a coiled-coil (CC) domain, is critical for Fusarium verticillioides virulence. In mammals, striatin interacts with multiple proteins to form a STRIPAK (striatin-interacting phosphatase and kinase) complex that regulates a variety of developmental processes and cellular mechanisms. In this study, we identified the homologue of a key mammalian STRIPAK component STRIP1/2 (striatin-interacting proteins 1 and 2) in F.verticillioides, FvStp1, which interacts with Fsr1 invivo. Gene deletion analysis indicates that FvStp1 is critical for F.verticillioides stalk rot virulence. In addition, we identified three proteins, designated FvCyp1, FvScp1 and FvSel1, which interact with the Fsr1 CC domain via a yeast two-hybrid screen. Importantly, FvCyp1, FvScp1 and FvSel1 co-localize to endomembrane structures, each having a preferred localization in the cell, and they are all required for F.verticillioides stalk rot virulence. Moreover, these proteins are necessary for the correct localization of Fsr1 to the endoplasmic reticulum (ER) and nuclear envelope. Thus, we identified several novel components in the STRIPAK complex that regulates F.verticillioides virulence, and propose that the correct organization and localization of Fsr1 are critical for STRIPAK complex function.

FvSTR1, a striatin orthologue in Fusarium virguliforme, is required for asexual development and virulence.

The soil-borne fungus Fusarium virguliforme causes sudden death syndrome (SDS), one of the most devastating diseases of soybean in North and South America. Despite the importance of SDS, a clear understanding of the fungal pathogenicity factors that affect the development of this disease is still lacking. We have identified FvSTR1, a F. virguliforme gene, which encodes a protein similar to a family of striatin proteins previously reported to regulate signalling pathways, cell differentiation, conidiation, sexual development, and virulence in filamentous fungi. Striatins are multi-domain proteins that serve as scaffolding units in the striatin-interacting phosphatase and kinase (STRIPAK) complex in fungi and animals. To address the function of a striatin homologue in F. virguliforme, FvSTR1 was disrupted and functionally characterized using a gene knock out strategy. The resulting Fvstr1 mutants were largely impaired in conidiation and pigmentation, and displayed defective conidia and conidiophore morphology compared to the wild-type and ectopic transformants. Greenhouse virulence assays revealed that the disruption of FvSTR1 resulted in complete loss of virulence in F. virguliforme. Microtome studies using fluorescence microscopy showed that the Fvstr1 mutants were defective in their ability to colonize the vascular system. The Fvstr1 mutants also showed a reduced transcript level of genes involved in asexual reproduction and in the production of secondary metabolites. These results suggest that FvSTR1 has a critical role in asexual development and virulence in F. virguliforme.

A STRIPAK complex mediates axonal transport of autophagosomes and dense core vesicles through PP2A regulation.

Autophagy plays an essential role in the cellular homeostasis of neurons, facilitating the clearance of cellular debris. This clearance process is orchestrated through the assembly, transport, and fusion of autophagosomes with lysosomes for degradation. The motor protein dynein drives autophagosome motility from distal sites of assembly to sites of lysosomal fusion. In this study, we identify the scaffold protein CKA (connector of kinase to AP-1) as essential for autophagosome transport in neurons. Together with other core components of the striatin-interacting phosphatase and kinase (STRIPAK) complex, we show that CKA associates with dynein and directly binds Atg8a, an autophagosomal protein. CKA is a regulatory subunit of PP2A, a component of the STRIPAK complex. We propose that the STRIPAK complex modulates dynein activity. Consistent with this hypothesis, we provide evidence that CKA facilitates axonal transport of dense core vesicles and autophagosomes in a PP2A-dependent fashion. In addition, CKA-deficient flies exhibit PP2A-dependent motor coordination defects. CKA function within the STRIPAK complex is crucial to prevent transport defects that may contribute to neurodegeneration.

An Epichloë festucae homologue of MOB3, a component of the STRIPAK complex, is required for the establishment of a mutualistic symbiotic interaction with Lolium perenne.

SummaryIn both Sordaria macrospora and Neurospora crassa, components of the conserved STRIPAK (striatin‐interacting phosphatase and kinase) complex regulate cell–cell fusion, hyphal network development and fruiting body formation. Interestingly, a number of Epichloë festucae genes that are required for hyphal cell–cell fusion, such as noxA, noxR, proA, mpkA and mkkA, are also required for the establishment of a mutualistic symbiotic interaction with Lolium perenne. To determine whether MobC, a homologue of the STRIPAK complex component MOB3 in S. macrospora and N. crassa, is required for E. festucae hyphal fusion and symbiosis, a mobC deletion strain was generated. The ΔmobC mutant showed reduced rates of hyphal cell–cell fusion, formed intrahyphal hyphae and exhibited enhanced conidiation. Plants infected with ΔmobC were severely stunted. Hyphae of ΔmobC showed a proliferative pattern of growth within the leaves of Lolium perenne with increased colonization of the intercellular spaces and vascular bundles. Although hyphae were still able to form expressoria, structures allowing the colonization of the leaf surface, the frequency of formation was significantly reduced. Collectively, these results show that the STRIPAK component MobC is required for the establishment of a mutualistic symbiotic association between E. festucae and L. perenne, and plays an accessory role in the regulation of hyphal cell–cell fusion and expressorium development in E. festucae.

The Strip-Hippo Pathway Regulates Synaptic Terminal Formation by Modulating Actin Organization at the Drosophila Neuromuscular Synapses

Synapse formation requires the precise coordination of axon elongation, cytoskeletal stability, and diverse modes of cell signaling. The underlying mechanisms of this interplay, however, remain unclear. Here, we demonstrate that Strip, a component of the striatin-interacting phosphatase and kinase (STRIPAK) complex that regulates these processes, is required to ensure the proper development of synaptic boutons at the Drosophila neuromuscular junction. In doing so, Strip negatively regulates the activity of the Hippo (Hpo) pathway, an evolutionarily conserved regulator of organ size whose role in synapse formation is currently unappreciated. Strip functions genetically with Enabled, an actin assembly/elongation factor and the presumptive downstream target of Hpo signaling, to modulate local actin organization at synaptic termini. This regulation occurs independently of the transcriptional co-activator Yorkie, the canonical downstream target of the Hpo pathway. Our study identifies a previously unanticipated role of the Strip-Hippo pathway in synaptic development, linking cell signaling to actin organization.

Catalytic Subunit 1 of Protein Phosphatase 2A Is a Subunit of the STRIPAK Complex and Governs Fungal Sexual Development.

ABSTRACTThe generation of complex three-dimensional structures is a key developmental step for most eukaryotic organisms. The details of the molecular machinery controlling this step remain to be determined. An excellent model system to study this general process is the generation of three-dimensional fruiting bodies in filamentous fungi like Sordaria macrospora. Fruiting body development is controlled by subunits of the highly conserved striatin-interacting phosphatase and kinase (STRIPAK) complex, which has been described in organisms ranging from yeasts to humans. The highly conserved heterotrimeric protein phosphatase PP2A is a subunit of STRIPAK. Here, catalytic subunit 1 of PP2A was functionally characterized. The Δpp2Ac1 strain is sterile, unable to undergo hyphal fusion, and devoid of ascogonial septation. Further, PP2Ac1, together with STRIPAK subunit PRO22, governs vegetative and stress-related growth. We revealed in vitro catalytic activity of wild-type PP2Ac1, and our in vivo analysis showed that inactive PP2Ac1 blocks the complementation of the sterile deletion strain. Tandem affinity purification, followed by mass spectrometry and yeast two-hybrid analysis, verified that PP2Ac1 is a subunit of STRIPAK. Further, these data indicate links between the STRIPAK complex and other developmental signaling pathways, implying the presence of a large interconnected signaling network that controls eukaryotic developmental processes. The insights gained in our study can be transferred to higher eukaryotes and will be important for understanding eukaryotic cellular development in general.

STRIPAK complexes in cell signaling and cancer.

Striatin-interacting phosphatase and kinase (STRIPAK) complexes are striatin-centered multicomponent supramolecular structures containing both kinases and phosphatases. STRIPAK complexes are evolutionarily conserved and have critical roles in protein (de)phosphorylation. Recent studies indicate that STRIPAK complexes are emerging mediators and regulators of multiple vital signaling pathways including Hippo, MAPK (mitogen-activated protein kinase), nuclear receptor and cytoskeleton remodeling. Different types of STRIPAK complexes are extensively involved in a variety of fundamental biological processes ranging from cell growth, differentiation, proliferation and apoptosis to metabolism, immune regulation and tumorigenesis. Growing evidence correlates dysregulation of STRIPAK complexes with human diseases including cancer. In this review, we summarize the current understanding of the assembly and functions of STRIPAK complexes, with a special focus on cell signaling and cancer.

A STRIPAK component Strip regulates neuronal morphogenesis by affecting microtubule stability.

During neural development, regulation of microtubule stability is essential for proper morphogenesis of neurons. Recently, the striatin-interacting phosphatase and kinase (STRIPAK) complex was revealed to be involved in diverse cellular processes. However, there is little evidence that STRIPAK components regulate microtubule dynamics, especially in vivo. Here, we show that one of the core STRIPAK components, Strip, is required for microtubule organization during neuronal morphogenesis. Knockdown of Strip causes a decrease in the level of acetylated α-tubulin in Drosophila S2 cells, suggesting that Strip influences the stability of microtubules. We also found that Strip physically and genetically interacts with tubulin folding cofactor D (TBCD), an essential regulator of α- and β-tubulin heterodimers. Furthermore, we demonstrate the genetic interaction between strip and Down syndrome cell adhesion molecule (Dscam), a cell surface molecule that is known to work with TBCD. Thus, we propose that Strip regulates neuronal morphogenesis by affecting microtubule stability.

The composition and function of the striatin-interacting phosphatases and kinases (STRIPAK) complex in fungi

The striatin-interacting phosphatases and kinases (STRIPAK) complex is a highly conserved eukaryotic protein complex that was recently described for diverse animal and fungal species. Here, we summarize our current knowledge about the composition and function of the STRIPAK complex from the ascomycete Sordaria macrospora, which we discovered by investigating sexually sterile mutants (pro), having a defect in fruiting body development. Mass spectrometry and yeast two-hybrid analysis defined core subunits of the STRIPAK complex, which have structural homologs in animal and other fungal organisms. These subunits (and their mammalian homologs) are PRO11 (striatin), PRO22 (STRIP1/2), SmMOB3 (Mob3), PRO45 (SLMAP), and PP2AA, the structural, and PP2Ac, the catalytic subunits of protein phosphatase 2A (PP2A). Beside fruiting body formation, the STRIPAK complex controls vegetative growth and hyphal fusion in S. macrospora. Although the contribution of single subunits to diverse cellular and developmental processes is not yet fully understood, functional analysis has already shown that mammalian homologs are able to substitute the function of distinct fungal STRIPAK subunits. This underscores the view that fungal model organisms serve as useful tools to get a molecular insight into cellular and developmental processes of eukaryotes in general. Future work will unravel the precise localization of single subunits within the cell and decipher their STRIPAK-related and STRIPAK-independent functions. Finally, evidence is accumulating that there is a crosstalk between STRIPAK and various signaling pathways, suggesting that eukaryotic development is dependent on STRIPAK signaling.

Germinal Center Kinases SmKIN3 and SmKIN24 Are Associated with the Sordaria macrospora Striatin-Interacting Phosphatase and Kinase (STRIPAK) Complex.

The striatin-interacting phosphatase and kinase (STRIPAK) complex is composed of striatin, protein phosphatase PP2A and protein kinases that regulate development in animals and fungi. In the filamentous ascomycete Sordaria macrospora, it is required for fruiting-body development and cell fusion. Here, we report on the presence and function of STRIPAK-associated kinases in ascomycetes. Using the mammalian germinal center kinases (GCKs) MST4, STK24, STK25 and MINK1 as query, we identified the two putative homologs SmKIN3 and SmKIN24 in S. macrospora. A BLASTP search revealed that both kinases are conserved among filamentous ascomycetes. The physical interaction of the striatin homolog PRO11 with SmKIN3 and SmKIN24 were verified by yeast two-hybrid (Y2H) interaction studies and for SmKIN3 by co-Immunoprecipitation (co-IP). In vivo localization found that both kinases were present at the septa and deletion of both Smkin3 and Smkin24 led to abnormal septum distribution. While deletion of Smkin3 caused larger distances between adjacent septa and increased aerial hyphae, deletion of Smkin24 led to closer spacing of septa and to sterility. Although phenotypically distinct, both kinases appear to function independently because the double-knockout strain ΔSmkin3/ΔSmkin24 displayed the combined phenotypes of each single-deletion strain.

Cell migration: changing shape.

Madsen et al. have delineated the roles of components of the striatin-interacting phosphatase and kinase (STRIPAK) complex in cancer cell migration and metastasis, which may explain why some of these proteins are overexpressed or mutated in cancer.

A fungal sarcolemmal membrane-associated protein (SLMAP) homolog plays a fundamental role in development and localizes to the nuclear envelope, endoplasmic reticulum, and mitochondria.

Sarcolemmal membrane-associated protein (SLMAP) is a tail-anchored protein involved in fundamental cellular processes, such as myoblast fusion, cell cycle progression, and chromosomal inheritance. Further, SLMAP misexpression is associated with endothelial dysfunctions in diabetes and cancer. SLMAP is part of the conserved striatin-interacting phosphatase and kinase (STRIPAK) complex required for specific signaling pathways in yeasts, filamentous fungi, insects, and mammals. In filamentous fungi, STRIPAK was initially discovered in Sordaria macrospora, a model system for fungal differentiation. Here, we functionally characterize the STRIPAK subunit PRO45, a homolog of human SLMAP. We show that PRO45 is required for sexual propagation and cell-to-cell fusion and that its forkhead-associated (FHA) domain is essential for these processes. Protein-protein interaction studies revealed that PRO45 binds to STRIPAK subunits PRO11 and SmMOB3, which are also required for sexual propagation. Superresolution structured-illumination microscopy (SIM) further established that PRO45 localizes to the nuclear envelope, endoplasmic reticulum, and mitochondria. SIM also showed that localization to the nuclear envelope requires STRIPAK subunits PRO11 and PRO22, whereas for mitochondria it does not. Taken together, our study provides important insights into fundamental roles of the fungal SLMAP homolog PRO45 and suggests STRIPAK-related and STRIPAK-unrelated functions.

Molecular modeling and molecular dynamics simulations based structural analysis of the SG2NA protein variants.

BackgroundSG2NA is a member of the striatin sub-family of WD-40 repeat proteins. Striatin family members have been associated with diverse physiological functions. SG2NA has also been shown to have roles in cell cycle progression, signal transduction etc. They have been known to interact with a number of proteins including Caveolin and Calmodulin and also propagate the formation of a multimeric protein unit called striatin-interacting phosphatase and kinase. As a pre-requisite for such interaction ability, these proteins are known to be unstable and primarily disordered in their arrangement. Earlier we had identified that it has multiple isoforms (namely 35, 78, 87 kDa based on its molecular weight) which are generated by alternative splicing. However, detailed structural information of SG2NA is still eluding the researchers.ResultsThis study was aimed towards three-dimensional molecular modeling and characterization of SG2NA protein and its isoforms. One structure out of five was selected for each variant having the least value for C score. Out of these, m35 kDa with a C score value of −3.21was the most poorly determined structure in comparison to m78 kDa and m87 kDa variants with C scores of −1.16 and −1.97 respectively. Further evaluation resulted in about 61.6% residues of m35 kDa, 76.6% residues of m78 kDa and 72.1% residues of m87 kDa falling in the favorable regions of Ramchandran Plot. Molecular dynamics simulations were also carried out to obtain biologically relevant structural models and compared with previous atomic coordinates. N-terminal region of all variants was found to be highly disordered.ConclusionThis study provides first-hand detailed information to understand the structural conformation of SG2NA protein variants (m35 kDa, m78 kDa and m87 kDa). The WD-40 repeat domain was found to constitute antiparallel strands of β-sheets arranged circularly. This study elucidates the crucial structural features of SG2NA proteins which are involved in various protein-protein interactions and also reveals the extent of disorder present in the SG2NA structure crucial for excessive interaction and multimeric protein complexes. The study also potentiates the role of computational approaches for preliminary examination of unknown proteins in the absence of experimental information.

Fam40b is required for lineage commitment of murine embryonic stem cells.

FAM40B (STRIP2) is a member of the striatin-interacting phosphatase and kinase (STRIPAK) complex that is involved in the regulation of various processes such as cell proliferation and differentiation. Its role for differentiation processes in embryonic stem cells (ESCs) is till now completely unknown. Short hairpin RNA (shRNA)-mediated silencing of Fam40b expression in ESCs and differentiating embryoid bodies (EBs) led to perturbed differentiation to embryonic germ layers and their derivatives including a complete abrogation of cardiomyogenesis. Pluripotency factors such as Nanog, Oct4 and Sox2 as well as epigenetic factors such as histone acetyltransferase type B (HAT1) and DNA (cytosine-5)-methyltransferase 3-β (Dnmt3b) were highly upregulated in Fam40b knockdown EBs as compared with control and scrambled EBs. To examine the relevance of Fam40b for development in vivo, Fam40b was knocked down in developing zebrafish. Morpholino-mediated knockdown of Fam40b led to severe abnormalities of the cardiovascular system, including an impaired expression of ventricular myosin heavy chain (vmhc) and of cardiac myosin light chain 2 (cmlc2) in the heart. We identified the gene product of Fam40b in ESCs as a perinuclear and nucleolar protein with a molecular weight of 96 kDa. We conclude that the expression of Fam40b is essential for the lineage commitment of murine embryonic stem cells (mESCs) into differentiated somatic cells via mechanisms involving pluripotency and epigenetic networks.

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

The protein phosphatase 2A (PP2A) and kinases such as germinal center kinase III (GCKIII) can interact with striatins to form a supramolecular complex called striatin-interacting phosphatase and kinase (STRIPAK) complex. Despite the fact that the STRIPAK complex regulates multiple cellular events, it remains only partially understood how this complex itself is assembled and regulated for differential biological functions. Our recent work revealed the activation mechanism of GCKIIIs by MO25, as well as how GCKIIIs heterodimerize with CCM3, a molecular bridge between GCKIII and striatins. Here we dissect the structural features of the coiled coil domain of striatin 3, a novel type of PP2A regulatory subunit that functions as a scaffold for the assembly of the STRIPAK complex. We have determined the crystal structure of a selenomethionine-labeled striatin 3 coiled coil domain, which shows it to assume a parallel dimeric but asymmetric conformation containing a large bend. This result combined with a number of biophysical analyses provide evidence that the coiled coil domain of striatin 3 and the PP2A A subunit form a stable core complex with a 2:2 stoichiometry. Structure-based mutational studies reveal that homodimerization of striatin 3 is essential for its interaction with PP2A and therefore assembly of the STRIPAK complex. Wild-type striatin 3 but not the mutants defective in PP2A binding strongly suppresses apoptosis of Jurkat cells induced by the GCKIII kinase MST3, most likely through a mechanism in which striatin recruits PP2A to negatively regulate the activation of MST3. Collectively, our work provides structural insights into the organization of the STRIPAK complex and will facilitate further functional studies.

Chapter Four - The Filamentous Fungus Sordaria macrospora as a Genetic Model to Study Fruiting Body Development

Filamentous fungi are excellent experimental systems due to their short life cycles as well as easy and safe manipulation in the laboratory. They form three-dimensional structures with numerous different cell types and have a long tradition as genetic model organisms used to unravel basic mechanisms underlying eukaryotic cell differentiation. The filamentous ascomycete Sordaria macrospora is a model system for sexual fruiting body (perithecia) formation. S. macrospora is homothallic, i.e., self-fertile, easily genetically tractable, and well suited for large-scale genomics, transcriptomics, and proteomics studies. Specific features of its life cycle and the availability of a developmental mutant library make it an excellent system for studying cellular differentiation at the molecular level. In this review, we focus on recent developments in identifying gene and protein regulatory networks governing perithecia formation. A number of tools have been developed to genetically analyze developmental mutants and dissect transcriptional profiles at different developmental stages. Protein interaction studies allowed us to identify a highly conserved eukaryotic multisubunit protein complex, the striatin-interacting phosphatase and kinase complex and its role in sexual development. We have further identified a number of proteins involved in chromatin remodeling and transcriptional regulation of fruiting body development. Furthermore, we review the involvement of metabolic processes from both primary and secondary metabolism, and the role of nutrient recycling by autophagy in perithecia formation. Our research has uncovered numerous players regulating multicellular development in S. macrospora. Future research will focus on mechanistically understanding how these players are orchestrated in this fungal model system.

SOcK, MiSTs, MASK and STicKs: the GCKIII (germinal centre kinase III) kinases and their heterologous protein–protein interactions

The GCKIII (germinal centre kinase III) subfamily of the mammalian Ste20 (sterile 20)-like group of serine/threonine protein kinases comprises SOK1 (Ste20-like/oxidant-stress-response kinase 1), MST3 (mammalian Ste20-like kinase 3) and MST4. Initially, GCKIIIs were considered in the contexts of the regulation of mitogen-activated protein kinase cascades and apoptosis. More recently, their participation in multiprotein heterocomplexes has become apparent. In the present review, we discuss the structure and phosphorylation of GCKIIIs and then focus on their interactions with other proteins. GCKIIIs possess a highly-conserved, structured catalytic domain at the N-terminus and a less-well conserved C-terminal regulatory domain. GCKIIIs are activated by tonic autophosphorylation of a T-loop threonine residue and their phosphorylation is regulated primarily through protein serine/threonine phosphatases [especially PP2A (protein phosphatase 2A)]. The GCKIII regulatory domains are highly disorganized, but can interact with more structured proteins, particularly the CCM3 (cerebral cavernous malformation 3)/PDCD10 (programmed cell death 10) protein. We explore the role(s) of GCKIIIs (and CCM3/PDCD10) in STRIPAK (striatin-interacting phosphatase and kinase) complexes and their association with the cis-Golgi protein GOLGA2 (golgin A2; GM130). Recently, an interaction of GCKIIIs with MO25 has been identified. This exhibits similarities to the STRADα (STE20-related kinase adaptor α)-MO25 interaction (as in the LKB1-STRADα-MO25 heterotrimer) and, at least for MST3, the interaction may be enhanced by cis-autophosphorylation of its regulatory domain. In these various heterocomplexes, GCKIIIs associate with the Golgi apparatus, the centrosome and the nucleus, as well as with focal adhesions and cell junctions, and are probably involved in cell migration, polarity and proliferation. Finally, we consider the association of GCKIIIs with a number of human diseases, particularly cerebral cavernous malformations.