Hyphal heterogeneity in <i><scp>A</scp>spergillus oryzae</i> is the result of dynamic closure of septa by <scp>W</scp>oronin bodies

Bleichrodt, Robert-Jan; Veluw, G. Jerre van; Recter, Brand; Maruyama, Jun‐ichi; Kitamoto, Katsuhiko; Wösten, Han A. B.

doi:10.1111/mmi.12077

Cited by 76 publications

(103 citation statements)

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“…Strain NSRKu70-1-1A, which was constructed by introducing plasmid pAdeA, containing the adeA marker gene, into strain NSRKu70-1-1 (18), was used as a wildtype strain. An Aohex1 disruptant strain, NSK-⌬hex1-1, was obtained by using strain NSRKu70-1-1, as previously described (13), and was used for comparative phenotype analysis in the same genetic background as that for the Aolah disruptant.…”

Section: Methodsmentioning

confidence: 99%

“…Although in a limited number of species, including N. crassa, the hexagonal crystal structure of the Woronin body is clearly visible by light microscopy, the fusion of Hex1 with a fluorescent protein is necessary to visualize and track Woronin bodies in most Pezizomycotina species (9,10). Such localization studies have revealed that Woronin bodies reversibly close the septal pore during normal growth and impede the cytoplasmic continuity between adjacent cells, thereby creating hyphal heterogeneity with regard to gene expression activity (13).…”

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confidence: 99%

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A Large Nonconserved Region of the Tethering Protein Leashin Is Involved in Regulating the Position, Movement, and Function of Woronin Bodies in Aspergillus oryzae

et al. 2014

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The Woronin body is a Pezizomycotina-specific organelle that is typically tethered to the septum, but upon hyphal wounding, it plugs the septal pore to prevent excessive cytoplasmic loss. Leashin (LAH) is a large Woronin body tethering protein that contains highly conserved N-and C-terminal regions and a long (ϳ2,500-amino-acid) nonconserved middle region. As the involvement of the nonconserved region in Woronin body function has not been investigated, here, we functionally characterized individual regions of the LAH protein of Aspergillus oryzae (AoLAH). In an Aolah disruptant, no Woronin bodies were tethered to the septum, and hyphae had a reduced ability to prevent excessive cytoplasmic loss upon hyphal wounding. Localization analysis revealed that the N-terminal region of AoLAH associated with Woronin bodies dependently on AoWSC, which is homologous to Neurospora crassa WSC (Woronin body sorting complex), and that the C-terminal region was localized to the septum. Elastic movement of Woronin bodies was observed when visualized with an AoLAH N-terminal-region-enhanced green fluorescent protein (EGFP) fusion protein. An N-and C-terminal fusion construct lacking the nonconserved middle region of AoLAH was sufficient for the tethering of Woronin bodies to the septum. However, Woronin bodies were located closer to the septum and exhibited impaired elastic movement. Moreover, expression of middle-region-deleted AoLAH in the Aolah disruptant did not restore the ability to prevent excessive cytoplasmic loss. These findings indicate that the nonconserved middle region of AoLAH has functional importance for regulating the position, movement, and function of Woronin bodies.

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Section: Methodsmentioning

confidence: 99%

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confidence: 99%

A Large Nonconserved Region of the Tethering Protein Leashin Is Involved in Regulating the Position, Movement, and Function of Woronin Bodies in Aspergillus oryzae

et al. 2014

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“…32) It is important to note that the septal pore is not constitutively open and that fluctuations between the open-closed state is dependent on the behavior of the AoSO protein in response to environmental stress. 33,34) The functional analysis of Woronin body brought a new aspect of biotin biosynthesis. Biotin is an essential cofactor involved in a number of carboxylation and decarboxylation reactions.…”

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confidence: 99%

Cell biology of the Koji mold Aspergillus oryzae

Kitamoto

2015

Bioscience, Biotechnology, and Biochemistry

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Koji mold, Aspergillus oryzae, has been used for the production of sake, miso, and soy sauce for more than one thousand years in Japan. Due to the importance, A. oryzae has been designated as the national micro-organism of Japan (Koku-kin). A. oryzae has been intensively studied in the past century, with most investigations focusing on breeding techniques and developing methods for Koji making for sake brewing. However, the understanding of fundamental biology of A. oryzae remains relatively limited compared with the yeast Saccharomyces cerevisiae. Therefore, we have focused on studying the cell biology including live cell imaging of organelles, protein vesicular trafficking, autophagy, and Woronin body functions using the available genomic information. In this review, I describe essential findings of cell biology of A. oryzae obtained in our study for a quarter of century. Understanding of the basic biology will be critical for not its biotechnological application, but also for an understanding of the fundamental biology of other filamentous fungi.

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“…Such zonal heterogeneity was also shown previously with respect to gene expression (38)(39)(40)(41)(42)(43)(44) and protein accumulation (39) in A. bisporus. In fact, these processes can be heterogeneous even between neighboring hyphae within a mycelial zone (45)(46)(47)(48)(49). Spectra of neighboring hyphae at the surface and under the peel of the mushroom were also heterogeneous (Fig.…”

Section: Figmentioning

confidence: 99%

Monitoring the Metabolic State of Fungal Hyphae and the Presence of Melanin by Nonlinear Spectral Imaging

Knaus

Blab

Agronskaia

et al. 2013

Appl Environ Microbiol

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b Label-free nonlinear spectral imaging microscopy (NLSM) records two-photon-excited fluorescence emission spectra of endogenous fluorophores within the specimen. Here, NLSM is introduced as a novel, minimally invasive method to analyze the metabolic state of fungal hyphae by monitoring the autofluorescence of NAD(P)H and flavin adenine dinucleotide (FAD). Moreover, the presence of melanin was analyzed by NLSM. NAD(P)H, FAD, and melanin were used as biomarkers for freshness of mushrooms of Agaricus bisporus (white button mushroom) that had been stored at 4°C for 0 to 17 days. During this period, the mushrooms did not show changes in morphology or color detectable by eye. In contrast, FAD/NAD(P)H and melanin/NAD(P)H ratios increased over time. For instance, these ratios increased from 0.92 to 2.02 and from 0.76 to 1.53, respectively, at the surface of mushroom caps that had been harvested by cutting the stem. These ratios were lower under the skin than at the surface of fresh mushrooms (0.78 versus 0.92 and 0.41 versus 0.76, respectively), indicative of higher metabolism and lower pigment formation within the fruiting body. Signals were different not only between tissues of the mushroom but also between neighboring hyphae. These data show that NLSM can be used to determine the freshness of mushrooms and to monitor the postharvest browning process at an early stage. Moreover, these data demonstrate the potential of NLSM to address a broad range of fundamental and applied microbiological processes. W orldwide, the annual production of edible mushrooms amounts to approximately 7.7 million tons. The white button mushroom, Agaricus bisporus, is one of the most important commercial species. It is low in fat and rich in fiber, vitamins, minerals, linoleic acid, and bioactive compounds such as anticancer polysaccharides (1-3). Moreover, the button mushroom contains more digestible proteins than most vegetables and only slightly less than meat products and milk (4, 5). Mushrooms are a perishable product. Therefore, there is a need for tools to monitor freshness of fruiting bodies. These tools could also be applied in breeding programs and to improve storage conditions.The metabolic state and melanin formation could be used as indicators for freshness of mushrooms. The presence of melanin can be detected by various microscopic techniques (6-8). There are also several methods to quantify the metabolic activity of fungi. They are often based on fluorescence assays monitoring the activity of NADH-dependent dehydrogenases (9) or NADH-dependent oxidoreductases (10, 11) or by viability stains that are metabolized in the presence of ATP and intracellular esterases (12). Most of these assays require cell extracts, tissue sectioning, and incorporation of marker molecules into cells. Nonlinear spectral microscopy (NLSM) is a potent, noninvasive (13-17) alternative to monitor the metabolic state and the presence of melanin in fungi. NLSM maps (auto)fluorescence emission spectra within the specimen (18)(19)(20). Phototoxicity of this ...

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Hyphal heterogeneity in Aspergillus oryzae is the result of dynamic closure of septa by Woronin bodies

Cited by 76 publications

References 59 publications

A Large Nonconserved Region of the Tethering Protein Leashin Is Involved in Regulating the Position, Movement, and Function of Woronin Bodies in Aspergillus oryzae

A Large Nonconserved Region of the Tethering Protein Leashin Is Involved in Regulating the Position, Movement, and Function of Woronin Bodies in Aspergillus oryzae

Cell biology of the Koji mold Aspergillus oryzae

Monitoring the Metabolic State of Fungal Hyphae and the Presence of Melanin by Nonlinear Spectral Imaging

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