scholarly journals Proliferation of Intrahyphal Hyphae Caused by Disruption ofcsmA, Which Encodes a Class V Chitin Synthase with a Myosin Motor-Like Domain in Aspergillus nidulans

1999 ◽  
Vol 181 (12) ◽  
pp. 3721-3729 ◽  
Author(s):  
Hiroyuki Horiuchi ◽  
Makoto Fujiwara ◽  
Shuichi Yamashita ◽  
Akinori Ohta ◽  
Masamichi Takagi
Keyword(s):  
Cell Wall ◽  
Aspergillus Nidulans ◽  
Wild Type Strain ◽  
Hyphal Growth ◽  
Chitin Synthase ◽  
Calcofluor White ◽  
Class V ◽  
Myosin Motor ◽  
Fungal Morphogenesis ◽  
Novel Protein

ABSTRACT We have found that the Aspergillus nidulans csmA gene encodes a novel protein which consists of an N-terminal myosin motor-like domain and a C-terminal chitin synthase domain (M. Fujiwara, H. Horiuchi, A. Ohta, and M. Takagi, Biochem. Biophys. Res. Commun. 236:75–78, 1997). To clarify the roles of csmA in fungal morphogenesis, we constructed csmA null mutants. The growth rate of the mutant colonies was almost the same as that of the wild-type strain, but hyphal growth was severely inhibited when a chitin-binding reagent, Calcofluor white or Congo red, was added to the medium. Moreover, morphological abnormalities in tip growth and septum formation were identified microscopically. Proliferation of intracellular new hyphae, called intrahyphal hyphae, which behaved as intrinsic hyphae, was the most striking phenotypic feature among them. These phenotypes were not suppressed when the only chitin synthase domain of csmA was expressed under the control of thealcA promoter, whereas they were suppressed when the intact form of csmA was expressed. Therefore, it was concluded that the product of csmA (CsmA) has important roles in polarized cell wall synthesis and maintenance of cell wall integrity and that the myosin motor-like domain is indispensable for these functions.

scholarly journals CsmA, a Class V Chitin Synthase with a Myosin Motor-like Domain, Is Localized through Direct Interaction with the Actin Cytoskeleton inAspergillus nidulans

2005 ◽  
Vol 16 (4) ◽  
pp. 1961-1970 ◽  
Author(s):  
Norio Takeshita ◽  
Akinori Ohta ◽  
Hiroyuki Horiuchi
Keyword(s):  
Cell Wall ◽  
Actin Cytoskeleton ◽  
Direct Interaction ◽  
Chitin Synthase ◽  
Structural Basis ◽  
Class V ◽  
Myosin Motor ◽  
Fungal Morphogenesis ◽  
Insight Into

One of the essential features of fungal morphogenesis is the polarized synthesis of cell wall components such as chitin. The actin cytoskeleton provides the structural basis for cell polarity in Aspergillus nidulans, as well as in most other eukaryotes. A class V chitin synthase, CsmA, which contains a myosin motor-like domain (MMD), is conserved among most filamentous fungi. The ΔcsmA null mutant showed remarkable abnormalities with respect to cell wall integrity and the establishment of polarity. In this study, we demonstrated that CsmA tagged with 9× HA epitopes localized near actin structures at the hyphal tips and septation sites and that its MMD was able to bind to actin. Characterization of mutants bearing a point mutation or deletion in the MMD suggests that the interaction between the MMD and actin is not only necessary for the proper localization of CsmA, but also for CsmA function. Thus, the finding of a direct interaction between the chitin synthase and the actin cytoskeleton provides new insight into the mechanisms of polarized cell wall synthesis and fungal morphogenesis.

scholarly journals Caspofungin Treatment of Aspergillus fumigatus Results in ChsG-Dependent Upregulation of Chitin Synthesis and the Formation of Chitin-Rich Microcolonies

2015 ◽  
Vol 59 (10) ◽  
pp. 5932-5941 ◽  
Author(s):  
Louise A. Walker ◽  
Keunsook K. Lee ◽  
Carol A. Munro ◽  
Neil A. R. Gow
Keyword(s):  
Cell Wall ◽  
Aspergillus Fumigatus ◽  
Dynamic Imaging ◽  
Chitin Synthase ◽  
Response To Treatment ◽  
Chitin Synthesis ◽  
Calcofluor White ◽  
Content Type ◽  
Chitin Content

ABSTRACTTreatment ofAspergillus fumigatuswith echinocandins such as caspofungin inhibits the synthesis of cell wall β-1,3-glucan, which triggers a compensatory stimulation of chitin synthesis. Activation of chitin synthesis can occur in response to sub-MICs of caspofungin and to CaCl2and calcofluor white (CFW), agonists of the protein kinase C (PKC), and Ca2+-calcineurin signaling pathways.A. fumigatusmutants with thechsgene (encoding chitin synthase) deleted (ΔAfchs) were tested for their response to these agonists to determine the chitin synthase enzymes that were required for the compensatory upregulation of chitin synthesis. Only the ΔAfchsGmutant was hypersensitive to caspofungin, and all other ΔAfchsmutants tested remained capable of increasing their chitin content in response to treatment with CaCl2and CFW and caspofungin. The resulting increase in cell wall chitin content correlated with reduced susceptibility to caspofungin in the wild type and all ΔAfchsmutants tested, with the exception of the ΔAfchsGmutant, which remained sensitive to caspofungin.In vitroexposure to the chitin synthase inhibitor, nikkomycin Z, along with caspofungin demonstrated synergistic efficacy that was againAfChsG dependent. Dynamic imaging using microfluidic perfusion chambers demonstrated that treatment with sub-MIC caspofungin resulted initially in hyphal tip lysis. However, thickened hyphae emerged that formed aberrant microcolonies in the continued presence of caspofungin. In addition, intrahyphal hyphae were formed in response to echinocandin treatment. Thesein vitrodata demonstrate thatA. fumigatushas the potential to survive echinocandin treatmentin vivobyAfChsG-dependent upregulation of chitin synthesis. Chitin-rich cells may, therefore, persist in human tissues and act as the focus for breakthrough infections.

scholarly journals Flumorph Is a Novel Fungicide That Disrupts Microfilament Organization in Phytophthora melonis

2007 ◽  
Vol 97 (5) ◽  
pp. 643-649 ◽  
Author(s):  
Shu Sheng Zhu ◽  
Xi Li Liu ◽  
Peng Fei Liu ◽  
Yong Li ◽  
Jian Qiang Li ◽  
...  
Keyword(s):  
Cell Wall ◽  
Tip Growth ◽  
Hyphal Growth ◽  
Characteristic Change ◽  
Calcofluor White ◽  
Actin Organization ◽  
Polarized Secretion ◽  
Phytophthora Melonis ◽  
Wall Materials ◽  
Cell Wall Materials

The mechanism of the effects of flumorph (a novel fungicide) was investigated by analyzing alterations of hyphal morphology, cell wall deposition patterns, F-actin organization, and other organelles in Phytophthora melonis. Calcofluor white staining suggested that flumorph did not inhibit the synthesis of cell wall materials, but disturbed the polar deposition of newly synthesized cell wall materials during cystospore germination and hyphal growth. After exposure to flumorph, zoospores were able to switch into cystospores accompanied with the formation of a cell wall, whereas cystospores failed to induce the isotropic-polar switch and did not produce germ tubes but continued the isotropic growth phase. In flumorph-treated hyphae, the most characteristic change was the development of periodic swelling (“beaded” morphology) and the disruption of tip growth. Newly synthesized cell wall materials were deposited uniformly throughout the diffuse expanded region of hyphae, in contrast to their normal polarized patterns of deposition. These alterations were the result of F-actin disruption, identified with the fluorescein isothiocynate (FITC)-phalloidin staining. The disruption of F-actin also was accompanied by disorganized organelles: each swelling of subapical hyphae was associated with a nucleus. Vesicles did not undergo polarized secretion to the apical hyphae, but diffused around nuclei for the subapical growth; thus, the cell wall was thickened with periodic expansion along the hyphae. Upon removing flumorph, normal tip growth and organized F-actin were observed again. These data, as well as data published earlier, suggest that flumorph may be involved in the impairment of cell polar growth through directly or indirectly disrupting the organization of F-actin. The primary site of action by flumorph in the disruption of the F-actin organization is under investigation.

scholarly journals Isolation ofcsm1encoding a class V chitin synthase with a myosin motor-like domain from the rice blast fungus,Pyricularia oryzae

1999 ◽  
Vol 170 (1) ◽  
pp. 131-139 ◽  
Author(s):  
In Cheol Park ◽  
Hiroyuki Horiuchi ◽  
Cher Won Hwang ◽  
Wan Hae Yeh ◽  
Akinori Ohta ◽  
...  
Keyword(s):  
Rice Blast ◽  
Rice Blast Fungus ◽  
Pyricularia Oryzae ◽  
Chitin Synthase ◽  
Class V ◽  
Myosin Motor ◽  
Blast Fungus

scholarly journals Aspergillus fumigatus RasA Regulates Asexual Development and Cell Wall Integrity

2008 ◽  
Vol 7 (9) ◽  
pp. 1530-1539 ◽  
Author(s):  
Jarrod R. Fortwendel ◽  
Kevin K. Fuller ◽  
Timothy J. Stephens ◽  
W. Clark Bacon ◽  
David S. Askew ◽  
...  
Keyword(s):  
Cell Wall ◽  
Hyphal Growth ◽  
Cell Wall Integrity ◽  
Signaling Cascades ◽  
Asexual Development ◽  
Wild Type ◽  
Calcofluor White ◽  
Ras Family ◽  
Type Gene ◽  
Increased Susceptibility

ABSTRACT The Ras family of proteins is a large group of monomeric GTPases. Members of the fungal Ras family act as molecular switches that transduce signals from the outside of the cell to signaling cascades inside the cell. A. fumigatus RasA is 94% identical to the essential RasA gene of Aspergillus nidulans and is the Ras family member sharing the highest identity to Ras homologs studied in many other fungi. In this study, we report that rasA is not essential in A. fumigatus, but its absence is associated with slowed germination and a severe defect in radial growth. The ΔrasA hyphae were more than two times the diameter of wild-type hyphae, and they displayed repeated changes in the axis of polarity during hyphal growth. The deformed hyphae accumulated numerous nuclei within each hyphal compartment. The ΔrasA mutant conidiated poorly, but this phenotype could be ameliorated by growth on osmotically stabilized media. The ΔrasA mutant also showed increased susceptibility to cell wall stressors, stained more intensely with calcofluor white, and was refractory to lysing enzymes used to make protoplasts, suggesting an alteration of the cell wall. All phenotypes associated with deletion of rasA could be corrected by reinsertion of the wild-type gene. These data demonstrate a crucial role for RasA in both hyphal growth and asexual development in A. fumigatus and provide evidence that RasA function is linked to cell wall integrity.

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