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Polarisome Meets Spitzenkörper: Microscopy, Genetics, and Genomics Converge

Plant Science Initiative, Department of Plant Pathology, University of Nebraska, Lincoln, NE 68588-0660, USA.
Eukaryotic Cell (Impact Factor: 3.18). 03/2005; 4(2):225-9. DOI: 10.1128/EC.4.2.225-229.2005
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    • "Endocytosis is thought to be important for conidial cells to detect certain external signals, and plays a crucial role in various aspects of hyphal tip growth (Fischer-Parton et al., 2000). Successful detection for the incorporation of FM4-64 into the Spitzenkörper body identified this organelle as being involved in endocytic membrane trafficking (Fischer-Parton et al., 2000; Harris et al., 2005; Verdin et al., 2009). Despite these findings, the physiological importance and molecular process of the endocytic process in filamentous fungi remain largely unaddressed, and little is known regarding the exocytotic transport of enzymes and other cell wall lytic materials in host tissue-invading processes (Gow et al., 2002). "
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    ABSTRACT: Endocytosis is an essential cellular process in eukaryotic cells that involves concordant functions of clathrin and adaptor proteins, various protein and lipid kinases, phosphatases and the actin cytoskeleton. In Saccharomyces cerevisiae, Ark1p is a member of the serine/threonine protein kinase (SPK) family that affects profoundly the organization of the cortical actin cytoskeleton. To study the function of MoArk1, an Ark1p homologue identified in Magnaporthe oryzae, we disrupted the MoARK1 gene and characterized the ΔMoark1 mutant strain. The ΔMoark1 mutant exhibited various defects ranging from mycelial growth and conidial formation to appressorium-mediated host infection. The ΔMoark1 mutant also exhibited decreased appressorium turgor pressure and attenuated virulence on rice and barley. In addition, the ΔMoark1 mutant displayed defects in endocytosis and formation of the Spitzenkörper, and was hyposensitive to exogenous oxidative stress. Moreover, a MoArk1-green fluorescent protein (MoArk1-GFP) fusion protein showed an actin-like localization pattern by localizing to the apical regions of hyphae. This pattern of localization appeared to be regulated by the N-ethylmaleimide-sensitive factor attachment protein receptor (SNARE) proteins MoSec22 and MoVam7. Finally, detailed analysis revealed that the proline-rich region within the MoArk1 serine/threonine kinase (S_TKc) domain was critical for endocytosis, subcellular localization and pathogenicity. These results collectively suggest that MoArk1 exhibits conserved functions in endocytosis and actin cytoskeleton organization, which may underlie growth, cell wall integrity and virulence of the fungus.
    Molecular Plant Pathology 02/2013; 14(5). DOI:10.1111/mpp.12020
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    • "This membrane-selective dye becomes incorporated into endocytic vesicle membranes and is then distributed to different organelles via the vesicle trafficking network via endosomes (Fisher-Parton et al. 2000; Read and Hickey 2001). In fungal hyphae, FM4-64 also stains vesicles within the Spk (Crampin et al. 2005; Dijksterhuis 2003; Fisher-Parton et al. 2000; Freitag et al. 2004; Hickey et al. 2005; Harris et al. 2005; Hoffman and Mendgen 1998; Peñalva 2005; Read and Hickey 2001; Torralba and Heath 2002; Riquelme et al. 2007; Köhli et al. 2008). "
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    ABSTRACT: Growing hyphae of Rhizoctonia solani were stained with the endocytic marker dye FM4-64 and imaged by confocal microscopy. Staining of the plasma membrane was followed by labeling of organelles in the cytoplasm (after ~1 min) and of the Spitzenkörper (Spk; after ~2 min). Fluorescence recovery after photobleaching (FRAP) of the stained Spk demonstrated the vectorial flow of secretory vesicles from the apical cytoplasm to the Spk. This flux was modelled in a two-compartment model. The turnover time of the vesicles of the Spk was estimated to be 1.3-2.5 min. These results are roughly consistent with the expected flux of vesicles through the Spk based on the number of secretory vesicles within the Spk and the number of secretory vesicles that would be necessary to fuse with the apical plasma membrane to maintain hyphal extension rates. These results suggest that membrane retrieval via endocytosis is not as significant as previously suggested.
    Antonie van Leeuwenhoek 01/2013; 103(4). DOI:10.1007/s10482-012-9873-1
    • "A key element involved in this transport equilibrium is the so-called Spitzenk€ orper that is thought to function as a vesicle supply center containing exocytotic as well as endocytotic vesicles (Virag and Harris 2006). The integrity of the Spitzenk€ orper depends on the polarisome, a protein complex that polarizes the actin-associated secretion machinery (Harris et al. 2005). Molecular components of the polarisome such the protein Spa2 (Carbo and Perez-Martin 2008) and supposedly formins, which nucleate actin cables, as well as components of the exocyst have been proposed to communicate with the Spitzenk€ orper (Harris 2009). "
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    ABSTRACT: Ustilago maydis is the causal agent of smut disease on corn plants. The infective process depends on the formation of a specific structure called infective filament consisting on a dikaryotic hyphae, which is required to penetrate the plant tissue. The formation of the infective filament in U. maydis is alike to a germination process, although it requires an intermediate mating step that links sexual development and virulence. This way, the induction of the pathogenic program implies strong morphological changes (bud to hypha transition) as well as genetic changes (haploid to dikaryotic transition). As a consequence, an accurate control of the cell cycle as well as morphogenesis is predicted during these transitions: the induction of the infective filament relies on a dual process that involves by one side a specific cell cycle arrest and in other side the specific activation of a hyperpolarization growth. Impairment of any of these processes will have as an outcome the inhibition of the virulence. This review has been framed in three major points: (1) Which transcriptional program is responsible for the induction of the infective filament formation, (2) How polar growth is regulated during the induction of the infective filament, and (3) Which mechanisms are responsible for cell cycle arrest during the infective filament formation.
    Morphogenesis and Pathogenicity in Fungi, 01/2012: pages 97-114;
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