Identification and Characterization of Dmct: A Cation Transporter in Yarrowia lipolytica Involved in Metal Tolerance.

Yarrowia lipolytica is a dimorphic fungus used as a model organism to investigate diverse biotechnological and biological processes, such as cell differentiation, heterologous protein production, and bioremediation strategies. However, little is known about the biological processes responsible for cation concentration homeostasis. Metals play pivotal roles in critical biochemical processes, and some are toxic at unbalanced intracellular concentrations. Membrane transport proteins control intracellular cation concentrations. Analysis of the Y. lipolytica genome revealed a characteristic functional domain of the cation efflux protein family, i.e., YALI0F19734g, which encodes YALI0F19734p (a putative Yl-Dmct protein), which is related to divalent metal cation tolerance. We report the in silico analysis of the putative Yl-Dmct protein's characteristics and the phenotypic response to divalent cations (Ca2+, Cu2+, Fe2+, and Zn2+) in the presence of mutant strains, Δdmct and Rdmct, constructed by deletion and reinsertion of the DMCT gene, respectively. The absence of the Yl-Dmct protein induces cellular and growth rate changes, as well as dimorphism differences, when calcium, copper, iron, and zinc are added to the cultured medium. Interestingly, the parental and mutant strains were able to internalize the ions. Our results suggest that the protein encoded by the DMCT gene is involved in cell development and cation homeostasis in Y. lipolytica.

Analysis of gene expression related to polyamine concentration and dimorphism induced in ornithine decarboxylase (odc) and spermidine synthase (spd) Ustilago maydis mutants.

Polyamines are ubiquitous small organic cations, and their roles as regulators of several cellular processes are widely recognized. They are implicated in the key stages of the fungal life cycle. Ustilago maydis is a phytopathogenic fungus, the causal agent of common smut of maize and a model system to understand dimorphism and virulence. U. maydis grows in yeast form at pH 7 and it can develop its mycelial form in vitro at pH 3. Δodc mutants that are unable to synthesize polyamines, grow as yeast at pH 3 with a low putrescine concentration, and to complete its dimorphic transition high putrescine concentration is require. Δspd mutants require spermidine to grow and cannot form mycelium at pH 3. In this work, the increased expression of the mating genes, mfa1 and mfa2, on Δodc mutants, was related to high putrescine concentration. Global gene expression analysis comparisons of Δodc and Δspd U. maydis mutants indicated that 2,959 genes were differentially expressed in the presence of exogenous putrescine at pH 7 and 475 genes at pH 3. While, in Δspd mutant, the expression of 1,426 genes was affected by exogenous spermine concentration at pH 7 and 11 genes at pH 3. Additionally, we identified 28 transcriptional modules with correlated expression during seven tested conditions: mutant genotype, morphology (yeast, and mycelium), pH, and putrescine or spermidine concentration. Furthermore, significant differences in transcript levels were noted for genes in modules relating to pH and genotype genes involved in ribosome biogenesis, mitochondrial oxidative phosphorylation, N-glycan synthesis, and Glycosylphosphatidylinositol (GPI)-anchor. In summary, our results offer a valuable tool for the identification of potential factors involved in phenomena related to polyamines and dimorphism.

Chitosan Is Necessary for the Structure of the Cell Wall, and Full Virulence of Ustilago maydis.

Smut fungi comprise a large group of biotrophic phytopathogens infecting important crops, such as wheat and corn. U. maydis is a plant pathogenic fungus responsible for common smut in maize and teocintle. Through our analysis of the transcriptome of the yeast-to-mycelium dimorphic transition at acid pH, we determined the number of genes encoding chitin deacetylases of the fungus, and observed that the gene encoding one of them (UMAG_11922; CDA1) was the only one up-regulated. The mutation of this gene and the analysis of the mutants revealed that they contained reduced amounts of chitosan, were severely affected in their virulence, and showed aberrant mycelial morphology when grown at acid pH. When the CDA1 gene was reinserted into the mutants by the use of an autonomous replication plasmid, virulence and chitosan levels were recovered in the retro mutant strains, indicating that the CDA1 gene was involved in these features. These data revealed that chitosan plays a crucial role in the structure and morphogenesis of the cell wall during mycelial development of the fungus, and that in its absence, the cell wall becomes altered and is unable to support the stress imposed by the defense mechanism mounted on by the plant host during the infection process.

Functions of polyamines in growth and development of Phycomyces blakesleeanus wild-type and mutant strains.

Polyamines are ubiquitous polycationic molecules with multiple effects. Spermidine was present in all the life stages of Phycomyces blakesleeanus, fulfilled the physiological requirement for polyamines during germination, and became most abundant at the emergence of germinating tubes. Putrescine was not found in resting spores or in stationary cultures, but was synthesized during apical growth and greatly exceeded spermidine in fast-growing stages of the vegetative and sexual life cycles. Changes in the polyamines did not correlate with the various stages of sporulation. Ornithine decarboxylase was so strongly inhibited in vitro by its product, putrescine, that it would completely block the enzyme if not compartmentalized away. 1,4-Diamino-2-butanone inhibited mycelial growth throughout the vegetative cycle without killing the cells. The inhibition was counteracted very effectively by putrescine, which acts as a close analog of the inhibitor, and very little by spermidine. Four independent spe mutants were obtained by a procedure that selected for resistance to diaminobutanone among functionally-uninucleate spores that survived exposure to N-methyl-N'-nitro-N-nitrosoguanidine. The stability of the enzyme, in vivo and in vitro, and its inhibition by diaminobutanone in vitro were the same in the wild type and in the mutants. Two of these were hypomorph mutants, with lower affinity of their ornithine decarboxylase for its substrate, ornithine, and lower maximal velocity. The other two were hypermorph transport mutants; we propose that they are affected in a protein that binds putrescine and its analogs for transport across the plasmalemma and sequestration away from the active enzyme. The transport mutants concentrated the exogenous diaminobutanone and the endogenous putrescine in inactive compartments; the highest enzyme activity was reached when the plasmalemma of the mutants was permeabilized with diethylaminoethyl dextran.

Tec1, a member of the TEA transcription factors family, is involved in virulence and basidiocarp development in Ustilago maydis / Tec1, un miembro de la familia de factores de transcripción TEA, está involucrado en la virulencia y el desarrollo de basidiocarpio en Ustilago maydis

The life cycle of Ustilago maydis involves alternation of a haploid saprophytic yeast-like stage and a dikaryotic hyphal virulent form. Under in vitro conditions, basidiocarps are formed. Analysis of the transcriptional network of basidiocarp formation revealed the possible involvement of a Tec transcription factor (Tec1, UMAG_02835) in the process. In some Ascomycota, Tec factors are involved in mycelial formation, pathogenesis, and interaction with other regulatory elements, but their role in Basidiomycota species is almost unknown. Accordingly, we proceeded to determine the role of this gene in U. maydis by its mutation. Tec1 was found to be a crucial factor for normal mating, basidiocarp development, and virulence, all of the functions related to the dikaryotic stage dependent of the b genes, whereas dimorphism and resistance to different stress conditions occurring in the haploid stage were not affected in tec1 mutants. The observation that mutants showed a low residual wild-type phenotype suggests the presence of a secondary mechanism that partially compensates the loss of Tec1.(AU)

Tec1, a member of the TEA transcription factors family, is involved in virulence and basidiocarp development in Ustilago maydis.

The life cycle of Ustilago maydis involves alternation of a haploid saprophytic yeast-like stage and a dikaryotic hyphal virulent form. Under in vitro conditions, basidiocarps are formed. Analysis of the transcriptional network of basidiocarp formation revealed the possible involvement of a Tec transcription factor (Tec1, UMAG_02835) in the process. In some Ascomycota, Tec factors are involved in mycelial formation, pathogenesis, and interaction with other regulatory elements, but their role in Basidiomycota species is almost unknown. Accordingly, we proceeded to determine the role of this gene in U. maydis by its mutation. Tec1 was found to be a crucial factor for normal mating, basidiocarp development, and virulence, all of the functions related to the dikaryotic stage dependent of the b genes, whereas dimorphism and resistance to different stress conditions occurring in the haploid stage were not affected in tec1 mutants. The observation that mutants showed a low residual wild-type phenotype suggests the presence of a secondary mechanism that partially compensates the loss of Tec1.

Identifying Genes Devoted to the Cell Death Process in the Gene Regulatory Network of Ustilago maydis.

Cell death is a process that can be divided into three morphological patterns: apoptosis, autophagy and necrosis. In fungi, cell death is induced in response to intracellular and extracellular perturbations, such as plant defense molecules, toxins and fungicides, among others. Ustilago maydis is a dimorphic fungus used as a model for pathogenic fungi of animals, including humans, and plants. Here, we reconstructed the transcriptional regulatory network of U. maydis, through homology inferences by using as templates the well-known gene regulatory networks (GRNs) of Saccharomyces cerevisiae, Aspergillus nidulans and Neurospora crassa. Based on this GRN, we identified transcription factors (TFs) as hubs and functional modules and calculated diverse topological metrics. In addition, we analyzed exhaustively the module related to cell death, with 60 TFs and 108 genes, where diverse cell proliferation, mating-type switching and meiosis, among other functions, were identified. To determine the role of some of these genes, we selected a set of 11 genes for expression analysis by qRT-PCR (sin3, rlm1, aif1, tdh3 [isoform A], tdh3 [isoform B], ald4, mca1, nuc1, tor1, ras1, and atg8) whose homologues in other fungi have been described as central in cell death. These genes were identified as downregulated at 72 h, in agreement with the beginning of the cell death process. Our results can serve as the basis for the study of transcriptional regulation, not only of the cell death process but also of all the cellular processes of U. maydis.

Acid pH Strategy Adaptation through NRG1 in Ustilago maydis.

The role of the Ustilago maydis putative homolog of the transcriptional repressor ScNRG1, previously described in Saccharomyces cerevisiae, Candida albicans and Cryptococcus neoformans, was analyzed by means of its mutation. In S. cerevisiae this gene regulates a set of stress-responsive genes, and in C. neoformans it is involved in pathogenesis. It was observed that the U. maydisNRG1 gene regulates several aspects of the cell response to acid pH, such as the production of mannosyl-erythritol lipids, inhibition of the expression of the siderophore cluster genes, filamentous growth, virulence and oxidative stress. A comparison of the gene expression pattern of the wild type strain versus the nrg1 mutant strain of the fungus, through RNA Seq analyses, showed that this transcriptional factor alters the expression of 368 genes when growing at acid pH (205 up-regulated, 163 down-regulated). The most relevant genes affected by NRG1 were those previously reported as the key ones for particular cellular stress responses, such as HOG1 for osmotic stress and RIM101 for alkaline pH. Four of the seven genes included WCO1 codifying PAS domain ( These has been shown as the key structural motif involved in protein-protein interactions of the circadian clock, and it is also a common motif found in signaling proteins, where it functions as a signaling sensor) domains sensors of blue light, two of the three previously reported to encode opsins, one vacuolar and non-pH-responsive, and another one whose role in the acid pH response was already known. It appears that all these light-reactive cell components are possibly involved in membrane potential equilibrium and as virulence sensors. Among previously described specific functions of this transcriptional regulator, it was found to be involved in glucose repression, metabolic adaptation to adverse conditions, cellular transport, cell rescue, defense and interaction with an acidic pH environment.

Wide distribution of the Ustilago maydis-bacterium endosymbiosis in naturally infected maize plants.

We have previously described that laboratory strains of Ustilago maydis, a fungal pathogen of maize and its ancestor teosinte, harbor an intracellular bacterium that enables the fungus to fix nitrogen. However, it is not clear whether other strains isolated from nature also harbor endosymbiotic bacteria, and whether these fix nitrogen for its host. In the present study, we isolated U. maydis strains from naturally infected maize. All the isolated strains harbored intracellular bacteria as determined by PCR amplification of the 16S rRNA gene, and some of them showed capacity to fix nitrogen. That these are truly bacterial endosymbionts were shown by the fact that, after thorough treatments with CuSO4 followed by serial incubations with antibiotics, the aforementioned bacterial gene was still amplified in treated fungi. In all, these data support the notion that U. maydis-bacterium endosymbiosis is a general phenomenon in this species.

Cell death in Ustilago maydis: comparison with other fungi and the effect of metformin and curcumin on its chronological lifespan.

Ustilago maydis is a Basidiomycota fungus, in which very little is known about its mechanisms of cell survival and death. To date, only the role of metacaspase1, acetate and hydrogen peroxide as inducers of cell death has been investigated. In the present work, we analyzed the lifespan of U. maydis compared with other species like Sporisorium reilianum, Saccharomyces cerevisiae and Yarrowia lipolytica, and we observed that U. maydis has a minor lifespan. We probe the addition of low concentrations metformin and curcumin to the culture media, and we observed that both prolonged the lifespan of U. maydis, a result observed for the first time in a phytopathogen fungus. However, higher concentrations of curcumin were toxic for the cells, and interestingly induced the yeast-to-mycelium dimorphic transition. The positive effect of metformin and curcumin appears to be related to an inhibition of the mechanistic Target of Rapamycin (mTOR) pathway, increase expression of autophagy genes and reducing of reactive oxygen species. These data indicate that U. maydis may be a eukaryotic model organism to elucidate the molecular mechanism underlying apoptotic and necrosis pathways, and the lifespan increase caused by metformin and curcumin.

The UMAG_00031 gene from Ustilago maydis encodes a putative membrane protein involved in pH control and morphogenesis.

We report the characterization of the gene UMAG_00031 from Ustilago maydis, previously identified as upregulated at alkaline pH. This gene is located on chromosome 1 and contains an ORF of 1539 bp that encodes a putative protein of 512 amino acids with an MW of 54.8 kDa. The protein is predicted to contain seven transmembrane domains (TMDs) and a signal peptide suggesting that is located in the cell membrane. Null ΔUMAG_00031 mutants were constructed, and their phenotype was analyzed. The mutant displayed a pleiotropic phenotype suggesting its participation in processes of alkaline pH adaptation independent of the Pal/Rim pathway. Also, it was involved in the dimorphic process induced by fatty acids. These results indicate that the protein encoded by the UMAG_00031 gene possibly functions as a receptor of different signals in the cell membrane of the fungus.

Parameters of Oxidative Stress in Reproductive and Postmenopausal Mexican Women.

In the reproductive phase, women experience cyclic changes in the ovaries and uterus, and hormones regulate these changes. Menopause is the permanent loss of menstruation after 12 months of amenorrhea. Menopause is also linked to a decrease in estrogen production, causing an imbalance in oxidative stress. We aimed to compare the three stages of lipid peroxidation, protein oxidative damage, and total antioxidant capacity (TAC) between reproductive-aged women (RAW) and postmenopausal women (PMW) in Mexico. We carried out a cross-sectional study with 84 women from Mexico City, including 40 RAW and 44 PMW. To determine the oxidative stress of the participants, several markers of lipid damage were measured: dienes conjugates (DC), lipohydroperoxides (LHP), and malondialdehyde (MDA); exposure to protein carbonyl is indicative of oxidative modified proteins, and TAC is indicative of the antioxidant defense system. Biomarkers of oxidative stress were significantly lower in RAW vs. PMW. DC were 1.31 ± 0.65 vs. 1.7 ± 0.51 pmol DC/mg dry weight (p = 0.0032); LHP were 4.95 ± 2.20 vs. 11.30 ± 4.24 pmol LHP/mg dry weight (p < 0.0001); malondialdehyde was 20.37 ± 8.20 vs. 26.10 ± 8.71 pmol MDA/mg dry weight (p = 0.0030); exposure of protein carbonyl was 3954 ± 884 vs. 4552 ± 1445 pmol PC/mg protein (p = 0.042); and TAC was 7244 ± 1512 vs. 8099 ± 1931 pmol Trolox equivalent/mg protein (p = 0.027). PMW display significantly higher oxidative stress markers compared to RAW; likewise, PMW show a higher TAC.

The signaling mechanisms involved in the dimorphic phenomenon of the Basidiomycota fungus Ustilago maydis.

In the present manuscript, we describe the mechanisms involved in the yeast-to-hypha dimorphic transition of the plant pathogenic Basidiomycota fungus Ustilago maydis. During its life cycle, U. maydis presents two stages: one in the form of haploid saprophytic yeasts that divide by budding and the other that is the product of the mating of sexually compatible yeast cells (sporidia), in the form of mycelial dikaryons that invade the plant host. The occurrence of the involved dimorphic transition is controlled by the two mating loci a and b. In addition, the dimorphic event can be obtained in vitro by different stimuli: change in the pH of the growth medium, use of different carbon sources, and by nitrogen depletion. The presence of other factors and mechanisms may affect this phenomenon; among these, we may cite the PKA and MAPK signal transduction pathways, polyamines, and factors that affect the structure of the nucleosomes. Some of these factors and conditions may affect all these dimorphic events, or they may be specific for only one or more but not all the processes involved. The conclusion reached by these experiments is that U. maydis has constituted a useful model for the analysis of the mechanisms involved in cell differentiation of fungi in general.

Multicellular growth of the Basidiomycota phytopathogen fungus Sporisorium reilianum induced by acid conditions.

Fungi are considered model organisms for the analysis of important phenomena of eukaryotes. For example, some of them have been described as models to understand the phenomenon of multicellularity acquisition by different unicellular organisms phylogenetically distant. Interestingly, in this work, we describe the multicellular development in the model fungus S. reilianum. We observed that Sporisorium reilianum, a Basidiomycota cereal pathogen that at neutral pH grows with a yeast-like morphology during its saprophytic haploid stage, when incubated at acid pH grew in the form of multicellular clusters. The multicellularity observed in S. reilianum was of clonal type, where buds of "stem" cells growing as yeasts remain joined by their cell wall septa, after cytokinesis. The elaboration and analysis of a regulatory network of S. reilianum showed that the putative zinc finger transcription factor CBQ73544.1 regulates a number of genes involved in cell cycle, cellular division, signal transduction pathways, and biogenesis of cell wall. Interestingly, homologous of these genes have been found to be regulated during Saccharomyces cerevisiae multicellular growth. In adddition, some of these genes were found to be negatively regulated during multicellularity of S. reilianum. With these data, we suggest that S. reilianum is an interesting model for the study of multicellular development.

Analysis of the photoreceptors involved in the light-depending basidiocarp formation in Ustilago maydis.

We have described that formation of basidiocarps by Ustilago maydis requires illumination. In the current research, we have proceeded to analyze what kind of light receptors are involved in this phenomenon. Accordingly, we investigated whether the homologues of the White Collar (WC), and the phytochrome (PHY) genes played a role in this process. Mutants deficient in either one of the three U. maydis WC homologue genes (WCO1a, WCO1b, WCO2), or the phytochrome-encoding the PHY gene were obtained. Phenotypic analysis of the mutants showed that ∆wco1a mutants formed similar numbers of basidiocarps than wild-type strain, whereas ∆wco1b mutants were severely affected in basidiocarp formation when illuminated with white, blue or red light. ∆wco2 and ∆phy1 mutants did not form basidiocarps under any illumination condition. These data indicate that Wco1a is the main blue light receptor, and Wco1b may operate as a secondary blue light receptor; Phy1 is the red light receptor, and Wco2 the transcription factor that controls the photo stimulation of the genes involved in the formation of fruiting bodies. It is suggested that effectiveness of the light receptors depends on the whole structure of the complex, possibly, because their association is necessary to maintain their functional structure.

Autophagosomes accumulation in the vacuoles of the fungus Ustilago maydis and the role of proteases in their digestion.

In the present study we determined whether Ustilago maydis accumulates autophagosomes within vacuoles when the cells are exposed to nutritional stress conditions. We investigated whether proteinase B and proteinase A are involved in their degradation. To this effect, wild type and Δpep4 mutant were incubated in minimal medium lacking a carbon source. It was observed that after incubation in nutrient-deficient media, spherical bodies appeared within the Δpep4 mutant strains vacuoles. In addition, autophagosomes were accumulated in U. maydis WT cells incubated in the presence of the serine protease inhibitor PMSF and accumulation of large autophagosomes and electrodense structures in the Δpep4 mutant cell vacuoles took place. These results demonstrate that the homologues of both, the proteinase B and the protease A, are involved in the autophagosomes degradation process in U. maydis.

Genotoxic effect of caffeine in Yarrowia lipolytica cells deficient in DNA repair mechanisms.

Caffeine is a compound that can exert physiological-beneficial effects in the organism. Nevertheless, there are controversies about its protective-antioxidant and/or its negative genotoxic effect. To abound on the analysis of the possible genotoxic/antioxidant effect of caffeine, we used as research model the yeast Yarrowia lipolytica parental strain, and mutant strains (∆rad52 and ∆ku80), which are deficient in the DNA repair mechanisms. Caffeine (5 mM) showed a cytostatic effect on all strains, but after 72 h of incubation the parental and ∆ku80 strains were able to recover of this inhibitory effect on growth, whereas ∆rad52 was unable to recover. When cells were pre-incubated with caffeine and H2O2 or incubated with a mixture of both agents, a higher inhibitory effect on growth of mutant strains was observed and this effect was noticeably greater for the Δrad52 strain. The toxic effect of caffeine appears to be through a mechanism of DNA damage (genotoxic effect) that involves DSB generation since, in all tested conditions, the growth of Δrad52 strain (cells deficient in HR DNA repair mechanism) was more severely affected.

Identification of a novel member of the pH responsive pathway Pal/Rim in Ustilago maydis.

The most important signal transduction mechanism related to environmental pH responses in fungi is the Pal/Rim pathway. Our knowledge of this pathway came initially from studies on Ascomycota species where it is made by seven members divided into two complexes, one located at the plasma membrane, and other at the endosomal membrane. In Basidiomycota sepecies only the homologs of the endosomal membrane complex (genes PalA/Rim20, PalB/ Rim13, and PalC/ Rim23), plus the transcription factor PacC/Rim101 have been identified. In this study, we describe the identification in Ustilago maydis of a gene encoding a Rho-like protein (tentatively named RHO4) as a novel member of this pathway. The RHO4 gene possibly plays, among other functions, a role in the second proteolytic cleavage that leads to the activation of the transcription factor PacC/Rim101. Mutants in this gene showed a pleiotropic phenotype, displaying similar characteristics to the Pal/Rim mutants, such as a lower growth rate at alkaline pH, high sensitivity to ionic and osmotic stresses, and impairment in protease secretion, but no alteration of the yeast-to-mycelium dimorphic transition induced by acid pH whereas it has a function in the dimorphic transition induced by fatty acids.

Cell wall glucans of fungi. A review.

Glucans are the most abundant polysaccharides in the cell walls of fungi, and their structures are highly variable. Accordingly, their glucose moieties may be joined through either or both alpha (α) or beta (ß) linkages, they are either lineal or branched, and amorphous or microfibrillar. Alpha 1,3 glucans sensu strictu (pseudonigerans) are the most abundant alpha glucans present in the cell walls of fungi, being restricted to dikarya. They exist in the form of structural microfibrils that provide resistance to the cell wall. The structure of beta glucans is more complex. They are linear or branched, and contain mostly ß 1,3 and ß 1,6 linkages, existing in the form of microfibrils. Together with chitin they constitute the most important structural components of fungal cell walls. They are the most abundant components of the cell walls in members of all fungal phyla, with the exception of Microsporidia, where they are absent. Taking into consideration the importance of glucans in the structure and physiology of the fungi, in the present review we describe the following aspects of these polysaccharides: i) types and distribution of fungal glucans, ii) their structure, iii) their roles, iv) the mechanism of synthesis of the most important ones, and v) the phylogentic relationships of the enzymes involved in their synthesis.

Analysis of the 3H8 antigen of Candida albicans reveals new aspects of the organization of fungal cell wall proteins.

The walls of both, yeast and mycelial cells of Candida albicans possess a species-specific antigen that is recognized by a monoclonal antibody (MAb 3H8). This antigen can be extracted in the form of a very high Mr complex, close or over 106 Da, by treatment, with ß-1,3-glucanase, ß mercaptoethanol or dithothreitol, or mild alkali, but not by saturated hydrogen fluoride (HF) in pyridine, suggesting that the complex is bound to wall ß-1,3 glucans, and to proteins by disulfide bonds, but not to ß-1,6 glucans. Through its sensitivity to trypsin and different deglycosylation procedures, it was concluded that the epitope is associated to a glycoprotein containing N-glycosidic, but not O-glycosidic mannan moieties. By means of electrophoresis in polycrylamide gradient gels, followed by mass spectrometric analysis, the epitope was pinpointed to a very high MW complex containing Agglutinin-Like Sequence (ALS) family proteins, and other cytoplasmic, membrane and secreted proteins. The components of this complex are bound by unknown covalent bonds. The material extracted with ß mercaptoethanol or dilute alkali appeared under the electron microscope as large aggregates in the form of spheroidal and mostly web-like structures of large sizes. These, and additional data, suggest that this protein complex may constitute an important part of the basic glycoprotein structure of C. albicans. The possibility that similar complexes exist in the wall of other fungi is an attractive, although yet untested possibility.