A seven-membered cell wall related transglycosylase gene family in Aspergillus niger is relevant for cell wall integrity in cell wall mutants with reduced α-glucan or galactomannan

Leeuwe, Tim M. van; Wattjes, Jasper; Niehues, Anna; Forn-Cuní, Gabriel; Geoffrion, Nicholas; Mélida, Hugo; Arentshorst, Mark; Molina, Antonio; Tsang, Adrian; Meijer, Annemarie H.; Moerschbacher, Bruno M.; Punt, Peter J.

doi:10.1016/j.tcsw.2020.100039

Cited by 16 publications

(12 citation statements)

References 90 publications

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“…The CRH genes of the GH16 family have been found in many other fungal genomes, including Aspergilli. However, in contrast to S. cerevisiae, systematic analyses of CRH mutants of A. fumigatus did not show increased susceptibility to CR and did not show any growth phenotypes (17,18). These results suggest that the function of CRH proteins may have been overemphasized in fungi and overlooked in cell wall construction.…”

Section: Discussionmentioning

confidence: 81%

“…A functional genomic study of Congo red activity in S. cerevisiae has allowed the identification of CRH (Congo red-hypersensitive) genes controlling susceptibility to Congo red (2). These genes do not, however, seem to play similar roles in filamentous fungi (17,18). Taken together, these data suggest that the effects of CR are not exclusively due to inhibition of structural cell wall polysaccharides and could involve multiple pathways (19)(20)(21).…”

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confidence: 99%

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Functional Genomic and Biochemical Analysis Reveals Pleiotropic Effect of Congo Red on Aspergillus fumigatus

Raj

Rhijn

et al. 2021

mBio

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Inhibition of fungal growth by Congo red (CR) has been putatively associated with specific binding to β-1,3-glucans, which blocks cell wall polysaccharide synthesis. In this study, we searched for transcription factors (TFs) that regulate the response to CR and interrogated their regulon. During the investigation of the susceptibility to CR of the TF mutant library, several CR-resistant and -hypersensitive mutants were discovered and further studied. Abnormal distorted swollen conidia called Quasimodo cells were seen in the presence of CR. Quasimodo cells in the resistant mutants were larger than the ones in the sensitive and parental strains; consequently, the conidia of the resistant mutants absorbed more CR than the germinating conidia of the sensitive or parental strains. Accordingly, this higher absorption rate by Quasimodo cells resulted in the removal of CR from the culture medium, allowing a subset of conidia to germinate and grow. In contrast, all resting conidia of the sensitive mutants and the parental strain were killed. This result indicated that the heterogeneity of the conidial population is essential to promote the survival of Aspergillus fumigatus in the presence of CR. Moreover, amorphous surface cell wall polysaccharides such as galactosaminogalactan control the influx of CR inside the cells and, accordingly, resistance to the drug. Finally, long-term incubation with CR led to the discovery of a new CR-induced growth effect, called drug-induced growth stimulation (DIGS), since the growth of one of them could be stimulated after recovery from CR stress. IMPORTANCE The compound Congo red (CR) has been historically used for coloring treatment and histological examination as well to inhibit the growth of yeast and filamentous fungi. It has been thought that CR binds to β-1,3-glucans in the fungal cell wall, disrupting the organization of the cell wall structure. However, other processes have been implicated in affecting CR sensitivity. Here, we explore CR susceptibility through screening a library of genetic null mutants. We find several previously uncharacterized genetic regulators important for CR susceptibility. Through biochemical and molecular characterization, we find cell membrane permeability to be important. Additionally, we characterize a novel cell type, Quasimodo cells, that occurs upon CR exposure. These cells take up CR, allowing the growth of the remaining fungi. Finally, we find that priming with CR can enhance long-term growth in one mutant.

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Section: Discussionmentioning

confidence: 81%

mentioning

confidence: 99%

Functional Genomic and Biochemical Analysis Reveals Pleiotropic Effect of Congo Red on Aspergillus fumigatus

Raj

Rhijn

et al. 2021

mBio

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“…Many possible cell wall synthesis or cell-wall-modifying enzymes could be targets of KexB processing. Explanations for the pH-dependent effect have been postulated and they suggest that at a lower pH, other proteases can perform KexB-like dibasic processing [28]. In yeast, yapsins, aspartic proteolytic enzymes involved in processing cell wall proteins including Scw4p, Utr2p, Pir4p and Gas1p [60][61][62], show a pH-dependent dependency on Kex2.…”

Section: Discussionmentioning

confidence: 99%

“…The deletion of kexB in A. niger is known to result in a pH-dependent morphological phenotype on solid medium [13]. On pH 6.0 buffered agar plates, the ∆kexB strain displays a hyper-branching and compact phenotype, whereas at pH 5.0 growth is like wild type [28]. To analyze the effect of the pH on the morphology during stirred submerged growth, we cultivated both the parental strain (N402) and ∆kexB at pH 5.0, 5.5 and 6.0.…”

Section: Disruption Of Kexb Shows a Ph-dependent Phenotype Between Ph 50 And Ph 60 In Fermentersmentioning

confidence: 99%

“…Using prediction algorithms such as v.1.0b ProPeptide Cleavage Site Prediction (ProP), a plethora of putative KexB targets can be identified, which are only indicative for potential targets and lack biological interpretation of any relation to the observed pleiotropic effects, such as shorter hyphae and a hyperbranching phenotype. Interestingly, this growth phenotype has clearly been shown on plates at pH 6.0, but not at pH 5.0, suggesting a pH-dependent phenotype [28], but a detailed study how the ∆kexB strain behaves during submerged growth is lacking. Here, we investigated the role of KexB on hyphal morphology in pH-controlled batch cultivations.…”

Section: Introductionmentioning

confidence: 98%

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Deletion of the Aspergillus niger Pro-Protein Processing Protease Gene kexB Results in a pH-Dependent Morphological Transition during Submerged Cultivations and Increases Cell Wall Chitin Content

et al. 2020

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There is a growing interest in the use of post-fermentation mycelial waste to obtain cell wall chitin as an added-value product. In the pursuit to identify suitable production strains that can be used for post-fermentation cell wall harvesting, we turned to an Aspergillus niger strain in which the kexB gene was deleted. Previous work has shown that the deletion of kexB causes hyper-branching and thicker cell walls, traits that may be beneficial for the reduction in fermentation viscosity and lysis. Hyper-branching of ∆kexB was previously found to be pH-dependent on solid medium at pH 6.0, but was absent at pH 5.0. This phenotype was reported to be less pronounced during submerged growth. Here, we show a series of controlled batch cultivations at a pH range of 5, 5.5, and 6 to examine the pellet phenotype of ΔkexB in liquid medium. Morphological analysis showed that ΔkexB formed wild type-like pellets at pH 5.0, whereas the hyper-branching ΔkexB phenotype was found at pH 6.0. The transition of phenotypic plasticity was found in cultivations at pH 5.5, seen as an intermediate phenotype. Analyzing the cell walls of ΔkexB from these controlled pH-conditions showed an increase in chitin content compared to the wild type across all three pH values. Surprisingly, the increase in chitin content was found to be irrespective of the hyper-branching morphology. Evidence for alterations in cell wall make-up are corroborated by transcriptional analysis that showed a significant cell wall stress response in addition to the upregulation of genes encoding other unrelated cell wall biosynthetic genes.

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Cephalosporin C biosynthesis and fermentation in Acremonium chrysogenum

Liu

Chen

Liu

et al. 2022

Appl Microbiol Biotechnol

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A seven-membered cell wall related transglycosylase gene family in Aspergillus niger is relevant for cell wall integrity in cell wall mutants with reduced α-glucan or galactomannan

Abstract: Highlights Aspergillus niger has seven chitin transglycosylation ( crh ) gene orthologues. All seven crh genes ( crhA-G ) are dispensable for growth and development. Disruption of crhA - G neither weakens the cell wall nor triggers the CWI response. … Show more

Cited by 16 publications

References 90 publications

Functional Genomic and Biochemical Analysis Reveals Pleiotropic Effect of Congo Red on Aspergillus fumigatus

Functional Genomic and Biochemical Analysis Reveals Pleiotropic Effect of Congo Red on Aspergillus fumigatus

Deletion of the Aspergillus niger Pro-Protein Processing Protease Gene kexB Results in a pH-Dependent Morphological Transition during Submerged Cultivations and Increases Cell Wall Chitin Content

Cephalosporin C biosynthesis and fermentation in Acremonium chrysogenum

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