Subcellular Compartmentalization and Trafficking of the Biosynthetic Machinery for Fungal Melanin

Upadhyay, Srijana; Xu, Xinping; Lowry, David; Jackson, Jennifer C.; Roberson, Robert W.; Lin, Xiaorong

doi:10.1016/j.celrep.2016.02.059

Cited by 77 publications

(87 citation statements)

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“…However, although some yeast species use classical secretory laccases to polymerize exogenously added precursors for melanization (14), the majority of filamentous fungi synthesize melanin de novo via the polyketide pathway (18). Like other secondary metabolism pathways, melanization through the polyketide pathway involves predicted cytosolic proteins, such as the polyketide synthase (PKS) and modification enzymes, as well as predicted conventional secretory laccases (19). For instance, the dihydroxynaphthalene (DHN) melanin biosynthesis pathway of Aspergillus fumigatus consists of six enzymes encoded by the melanin gene cluster (Fig.…”

Section: Introductionmentioning

confidence: 99%

“…Based on the order of reactions that they carry out, the polyketide synthase Alb1 (also known as PksP) and the modification enzymes Ayg1, Arp1, and Arp2 are categorized as early enzymes, whereas the two laccases Abr1 and Abr2 are categorized as late enzymes (19). We demonstrated recently that the two late enzymes, laccases Abr1 and Abr2, are indeed secretory proteins and that they accumulate in the cell wall of conidiophores and conidia (19, 20). Surprisingly, contrary to the predicted cytoplasmic localization, all four early melanin enzymes are localized to the secretory endosomes (19).…”

Section: Introductionmentioning

confidence: 99%

“…We demonstrated recently that the two late enzymes, laccases Abr1 and Abr2, are indeed secretory proteins and that they accumulate in the cell wall of conidiophores and conidia (19, 20). Surprisingly, contrary to the predicted cytoplasmic localization, all four early melanin enzymes are localized to the secretory endosomes (19). The discovery of all melanin enzymes trafficking to/through endosomes provides a plausible explanation for the subcellular compartmentalization of melanin biosynthesis and trafficking in secretory endosomes in Aspergillus .…”

Section: Introductionmentioning

confidence: 99%

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Interactions between Melanin Enzymes and Their Atypical Recruitment to the Secretory Pathway by Palmitoylation

Upadhyay

Lin

2016

mBio

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Melanins are biopolymers that confer coloration and protection to the host organism against biotic or abiotic insults. The level of protection offered by melanin depends on its biosynthesis and its subcellular localization. Previously, we discovered that Aspergillus fumigatus compartmentalizes melanization in endosomes by recruiting all melanin enzymes to the secretory pathway. Surprisingly, although two laccases involved in the late steps of melanization are conventional secretory proteins, the four enzymes involved in the early steps of melanization lack a signal peptide or a transmembrane domain and are thus considered “atypical” secretory proteins. In this work, we found interactions among melanin enzymes and all melanin enzymes formed protein complexes. Surprisingly, the formation of protein complexes by melanin enzymes was not critical for their trafficking to the endosomal system. By palmitoylation profiling and biochemical analyses, we discovered that all four early melanin enzymes were strongly palmitoylated during conidiation. However, only the polyketide synthase (PKS) Alb1 was strongly palmitoylated during both vegetative hyphal growth and conidiation when constitutively expressed alone. This posttranslational lipid modification correlates the endosomal localization of all early melanin enzymes. Intriguingly, bioinformatic analyses predict that palmitoylation is a common mechanism for potential membrane association of polyketide synthases (PKSs) and nonribosomal peptide synthetases (NRPSs) in A. fumigatus. Our findings indicate that protein-protein interactions facilitate melanization by metabolic channeling, while posttranslational lipid modifications help recruit the atypical enzymes to the secretory pathway, which is critical for compartmentalization of secondary metabolism.

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Section: Introductionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

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Interactions between Melanin Enzymes and Their Atypical Recruitment to the Secretory Pathway by Palmitoylation

Upadhyay

Lin

2016

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“…Thus, carbon flow during cyclosporine biosynthesis seems to be directed from the cytosol to the vacuolar lumen and during penicillin biosynthesis from the vacuole to the cytosol to peroxisomes (reviewed in Roze et al, 2011), whereas metabolite channelling seems to be directed from the cytosol to the ER in F. graminearum. In addition, the biosynthesis of another fungal secondary metabolite melanin is also compartmentalized (Upadhyay et al, 2016). In Aspergillus fumigatus and A. nidulans cytosolic enzymes involved in early steps of melanin biosynthesis are recruited to endosomes through a non-conventional secretory pathway, whereas late melanin enzymes accumulate in the cell wall (Upadhyay et al, 2016).…”

Section: Is Subcellular Partitioning a General Phenomenon In Fungal Smentioning

confidence: 99%

“…In addition, the biosynthesis of another fungal secondary metabolite melanin is also compartmentalized (Upadhyay et al, 2016). In Aspergillus fumigatus and A. nidulans cytosolic enzymes involved in early steps of melanin biosynthesis are recruited to endosomes through a non-conventional secretory pathway, whereas late melanin enzymes accumulate in the cell wall (Upadhyay et al, 2016).…”

Section: Is Subcellular Partitioning a General Phenomenon In Fungal Smentioning

confidence: 99%

Regulation and cell biology of secondary metabolite production in Fusarium graminearum and Fusarium pseudograminearum

Blum¹

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1 AbstractAscomycete fungi have the potential to produce a vast array of secondary metabolites, which are metabolites not required for normal growth and development. In any one species there is typically the potential for production of upward of fifty of these compounds. Many fungal secondary metabolites are used as medicines (e.g. penicillin or cyclosporine), or are of importance due to their toxicity in humans or animals (e.g. aflatoxins), or because they are virulence factors during crop plant infection (e.g. deoxynivalenol). Despite the vast potential encoded in ascomycete genomes, the vast majority of fungal secondary metabolites are yet to be discovered. This is in part due to our lack of understanding of how the production of these compounds is regulated. Moreover, even for the compounds we do know, we are just beginning to understand the complex interaction of external stimuli, signalling pathways, cellular and developmental biology that leads to their production. In this thesis, biological insights into these aspects of two fungal secondary metabolites, deoxynivalenol (DON) and fusatin are provided.In chapter II a high-throughput fluorescence activated cell sorting (FACS) based mutant screen was developed and the regulation of DON production in Fusarium graminearum was analysed. DON is an economically very important mycotoxin, as it is the most common contaminant of wheat, barley and corn worldwide. It is produced by plant pathogenic filamentous fungi of the Fusarium family, which can cause Fusarium head blight as well as Fusarium crown rot. During plant infection, DON is an important virulence factor and it can be toxic for humans and animals. The regulation of DON production is not yet fully understood. To identify regulators, a mutant screen was developed using FACS to allow high throughput screening, coupled with whole genome sequencing to identify mutations.Segregation analyses were performed to determine the mutation causing the phenotype.The mutant screen identified an adenylyl cyclase allele, which resulted in production of DON under normally repressive conditions. In addition, the mutant developed cellular structures associated with toxin production under repressive conditions. The levels of the fundamental second messenger cAMP, which adenylyl cyclases produce, were increased in the identified mutant, suggesting it might be a gain-of-function adenylyl cyclase mutant. This mutant screening methodology can be adapted to identify regulators of different secondary metabolites as well as to identify mutants overproducing yet unknown secondary metabolites. Abstract 2In chapter III the cell biology of DON production in Fusarium graminearum was elucidated with quantitative super resolution microscopy. During the last five years the first insights into the cell biology of DON production were gained, indicating complex cell biological mechanisms. During toxin production, F. graminearum develops endoplasmic reticulum proliferations, called toxisomes, at which two DON biosynthesis enzymes, TRI1 a...

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Autolytic enzymes are responsible for increased melanization of carbon stressed Aspergillus nidulans cultures

et al. 2017

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Melanization of carbon stressed Aspergillus nidulans cultures were studied. Melanin production showed strong positive correlation with the activity of the secreted chitinase and ß-1,3-glucanase. Deletion of either chiB encoding an autolytic endochitinase or engA encoding an autolytic ß-1,3-endoglucanase, or both, almost completely prevented melanization of carbon stressed cultures. In contrast, addition of Trichoderma lyticase to cultures induced melanin production. Synthetic melanin could efficiently inhibit the purified ChiB chitinase activity. It could also efficiently decrease the intensity of hyphal fragmentation and pellet disorganization in Trichoderma lyticase treated cultures. Glyphosate, an inhibitor of L-3,4-dihydroxyphenylalanine-type melanin synthesis, could prevent melanization of carbon-starved cultures and enhanced pellet disorganization, while pyroquilon, a 1,8-dihydroxynaphthalene-type melanin synthesis inhibitor, enhanced melanization, and prevented pellet disorganization. We concluded that cell wall stress induced by autolytic cell wall hydrolases was responsible for melanization of carbon-starved cultures. The produced melanin can shield the living cells but may not inhibit the degradation and reutilization of cell wall materials of dead hyphae. Controlling the activity of autolytic hydrolase production can be an efficient approach to prevent unwanted melanization in the fermentation industry, while applying melanin synthesis inhibitors can decrease the resistance of pathogenic fungi against the chitinases produced by the host organism.

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Subcellular Compartmentalization and Trafficking of the Biosynthetic Machinery for Fungal Melanin

Cited by 77 publications

References 47 publications

Interactions between Melanin Enzymes and Their Atypical Recruitment to the Secretory Pathway by Palmitoylation

Interactions between Melanin Enzymes and Their Atypical Recruitment to the Secretory Pathway by Palmitoylation

Regulation and cell biology of secondary metabolite production in Fusarium graminearum and Fusarium pseudograminearum

Autolytic enzymes are responsible for increased melanization of carbon stressed Aspergillus nidulans cultures

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