PubMed Health⌕ Search

Biomedical subjects

Michelle Momany

Publications and source records attributed to Michelle Momany.

At least 19 recordsLinked to original sources
PubMed2007

The first fifty microarray studies in filamentous fungi.

Microarray studies have examined global gene expression in over 20 species of filamentous fungi encompassing a wide variety of research areas. The majority have addressed aspects of metabolism or pathogenicity. Metabolic studies have revealed important differences in the transcriptional regulation of genes for primary metabolic pathways between filamentous fungi and yeast. Transcriptional profiles for genes involved in secondary metabolism have also been established. Genes required for the biosynthesis of both useful and detrimental secondary metabolites have been identified. Due to the economic, ecological and medical implications, it is not surprising that many studies have used microarray analysis to examine gene expression in pathogenic filamentous fungi. Genes involved in various stages of pathogenicity have been identified, including those thought to be important for adaptation to the host environment. While most of the studies have simulated pathogenic conditions in vitro, a small number have also reported fungal gene expression within their plant hosts. This review summarizes the first 50 microarray studies in filamentous fungi and highlights areas for future investigation.

Adaptation, Physiological↗
PubMed2006

Septin localization across kingdoms: three themes with variations.

Septins are GTPases that form filaments in fungi and animals. In addition to their original role in cell division, septins have been shown to have roles in coordinating nuclear division, membrane trafficking and organizing the cytoskeleton. Many recent studies have examined subcellular localization of septins in a wide range of fungi and animals. Septin localization shows three patterns, which generally correspond to function across kingdoms. Septins that localize to projections shape and compartmentalize emerging growth. Septins that localize to partitions compartmentalize pre-existing cellular material. Septins that localize to the whole cell are involved in membrane trafficking and organizing the cytoskeleton and are most often in animals. The difference in localization pattern frequency between kingdoms will probably disappear as more septins are examined in diverse organisms and tissues.

Animals↗
PubMed2006

FsFKS1, the 1,3-beta-glucan synthase from the caspofungin-resistant fungus Fusarium solani.

The cell wall, a mesh of carbohydrates and proteins, shapes and protects the fungal cell. The enzyme responsible for the synthesis of one of the main components of the fungal wall, 1,3-beta-glucan synthase, is targeted by the antifungal caspofungin acetate (CFA). Clinical isolates of Candida albicans and Aspergillus fumigatus are much more sensitive to CFA than clinical isolates of Fusarium species. To better understand CFA resistance in Fusarium species, we cloned and sequenced FsFKS1, which encodes the Fusarium solani f. sp. pisi beta(1,3)-D-glucan synthase, used RNA interference to reduce its expression and complemented deletion of the essential fks gene of the CFA-sensitive fungus A. fumigatus with FsFKS1. Reduction of the FsFKS1 message in F. solani f. sp. pisi reduced spore viability and caused lysis of spores and hyphae, consistent with cell wall defects. Compensating for the loss of A. fumigatus fks1 with FsFKS1 caused only a modest increase in the tolerance of A. fumigatus for CFA. Our results suggest that FsFKS1 is required for the proper construction of F. solani cell walls and that the resistance of F. solani to CFA is at best only partially due to resistance of the FsFKS1 enzyme to this antifungal agent.

Caspofungin↗
PubMed2005

Sequencing of Aspergillus nidulans and comparative analysis with A. fumigatus and A. oryzae.

The aspergilli comprise a diverse group of filamentous fungi spanning over 200 million years of evolution. Here we report the genome sequence of the model organism Aspergillus nidulans, and a comparative study with Aspergillus fumigatus, a serious human pathogen, and Aspergillus oryzae, used in the production of sake, miso and soy sauce. Our analysis of genome structure provided a quantitative evaluation of forces driving long-term eukaryotic genome evolution. It also led to an experimentally validated model of mating-type locus evolution, suggesting the potential for sexual reproduction in A. fumigatus and A. oryzae. Our analysis of sequence conservation revealed over 5,000 non-coding regions actively conserved across all three species. Within these regions, we identified potential functional elements including a previously uncharacterized TPP riboswitch and motifs suggesting regulation in filamentous fungi by Puf family genes. We further obtained comparative and experimental evidence indicating widespread translational regulation by upstream open reading frames. These results enhance our understanding of these widely studied fungi as well as provide new insight into eukaryotic genome evolution and gene regulation.

Aspergillus fumigatus↗
PubMed2004

Aspergillus nidulans RhoA is involved in polar growth, branching, and cell wall synthesis.

Growth of the filamentous fungus Aspergillus nidulans begins when the conidium breaks dormancy and grows isotropically. Eventually a germ tube emerges and the axis of growth remains fixed in the primary hypha while new growth axes are established basally to form secondary germ tubes and lateral branches. Rho1 is a Rho family GTPase that has been shown to be involved in polarity establishment and cell wall deposition in Saccharomyces cerevisiae. A gene predicted to encode a Rho1 homolog was cloned from A. nidulans and named rhoA. Strains carrying ectopic copies of the constitutively active rhoA(G14V) allele or the dominant rhoA(E40I) allele were created and characterized. The constitutively active rhoA(G14V) strain grew slowly relative to wild type and showed an abnormal clustered pattern of branch emergence. The rhoA(G14V) strain also labeled intensely with calcofluor, showed elevated levels of cell wall N-acetylglucosamine and had unusually thick cell walls. The dominant rhoA(E40I)strain was accelerated in the emergence of secondary and tertiary germ tubes, and lateral branches relative to wild type and showed lysis with prolonged incubation. The rhoA(E40I) strain also was hypersensitive to the cell wall disrupting agents calcofluor and caspofungin acetate and showed an increase in cell wall N-acetylglucosamine levels. Our results suggest that rhoA plays a role in polarity, proper branching pattern, and cell wall deposition.

Amino Acid Sequence↗
PubMed2004

Polarity in filamentous fungi: moving beyond the yeast paradigm.

Filamentous fungi grow by the polar extension of hyphae. This polar growth requires the specification of sites of germ tube or branch emergence, followed by the recruitment of the morphogenetic machinery to those sites for localized cell wall deposition. Researchers attempting to understand hyphal morphogenesis have relied upon the powerful paradigm of bud emergence in the yeast Saccharomyces cerevisiae. The yeast paradigm has provided a useful framework, however several features of hyphal morphogenesis, such as the ability to maintain multiple axes of polarity and an extremely rapid extension rate, cannot be explained by simple extrapolation from yeast models. We discuss recent polarity research from filamentous fungi focusing on the position of germ tube emergence, the relaying of positional information via RhoGTPase modules, and the recruitment of morphogenetic machinery components including cytoskeleton, polarisome and ARP2/3 complexes, and the vesicle trafficking system.

Cell Polarity↗
PubMed2004

Identification and complementation of abnormal hyphal branch mutants ahbA1 and ahbB1 in Aspergillus nidulans.

Branching generates new axes of polar growth in filamentous fungi and is critical for development, reproduction, and pathogenicity. To investigate branching we screened an Aspergillus nidulans temperature-sensitive mutant collection for abnormal hyphal branch (ahb) mutants. We identified two mutants, ahbA1, which showed reduced branching relative to wild type at restrictive temperature, and ahbB1, which showed increased branching relative to wild type at restrictive temperature. Both mutants also showed abnormal conidiophore development at restrictive temperature. The ahbA1 hypobranching mutant showed defects in nuclear division and hydroxyurea resistance. Complementation and sequencing showed that ahbA1 is a previously identified allele of the cell cycle regulator nimX. The ahbB1 hyperbranching mutant had an increased number of nuclei, was osmotically remedial and Calcofluor resistant. The ahbB gene is predicted to encode a novel protein that has homologues exclusively in filamentous fungi. The C-terminal domain of the predicted AhbB protein showed homology with the heme-binding domain of a cytochrome P450 protein and sequencing of the ahbB1 mutant allele showed that the lesion lies just before this putative heme-binding domain. The ahbB1 mutant showed increased sensitivity to the ergosterol biosynthesis inhibitor imidazole. Our results suggest a link between nuclear division and branching and a possible role for membrane synthesis in branching.

Amino Acid Sequence↗
PubMed2004

The Aspergillus fumigatus cell wall is organized in domains that are remodelled during polarity establishment.

Aspergillus fumigatus is a life-threatening and increasingly frequent pathogen of the immunocompromised. Like other filamentous fungi A. fumigatus grows in a highly polar manner, adding new cell wall to the apical region of hyphae. mAbs were raised against isolated A. fumigatus cell walls. Fifteen antibodies bound reproducibly to isolated cell walls in ELISAs and to the walls of intact cells in immunofluorescence experiments. Surprisingly, individual mAbs showed distinct patterns of localization. Six antibodies labelled exclusively conidial or basal regions, seven labelled apical regions and a single antibody labelled both basal and apical regions of hyphae. Ten antibodies did not label the walls adjacent to septa. In double labelling experiments with representative mAbs there was little or no overlap between epitopes recognized. These labelling patterns suggest that the wall is made up of basal and apical domains that differ in composition or organization and that the wall region flanking septa differs from other regions of the lateral wall. In time-course experiments of early A. fumigatus growth, mAb16C4 failed to label isotropically expanding cells and labelled emerging germ tubes and branches. The same mAb failed to label the Aspergillus nidulans swoC mutant, which is defective in polarity establishment. However, mAb16C4 did label the A. nidulans swoA mutant, which completes polarity establishment, but is defective in polarity maintenance. Thus, mAb16C4 appears to recognize a cell wall change that occurs during polarity establishment. In immunogold labelling and transmission electron microscopy (TEM) experiments, conidia, basal regions and apical regions of thin-sectioned cells labelled with mAb16C4. That only apical regions labelled in intact cells (immunofluorescence) while conidial, basal and apical regions labelled in thin-sectioned cells (TEM) suggests that the 16C4 epitope is present along the whole hypha, but is masked everywhere except the tip until polarity establishment. That is to say, some remodelling of the wall during polarity establishment exposes the 16C4 epitope. The 16C4 epitope was partially purified from A. fumigatus total protein by passage through hydrophobic interaction and anion-exchange columns. The resulting single ELISA-positive fraction showed relatively few bands by SDS-PAGE and silver staining and a strong band by Western blotting with the16C4 mAb. Sequencing of the fraction yielded a predicted peptide with a 6-amino acid exact match to a region of the Cat1 protein previously identified as an immunodominant A. fumigatus catalase that localizes to the cell wall and is secreted to the medium. Experiments are under way to determine if mAb16C4 recognizes Cat1 or another protein that co-purifies with Cat1.

Antigens, Fungal↗
PubMed2003

Glycosphingolipids of the model fungus Aspergillus nidulans: characterization of GIPCs with oligo-alpha-mannose-type glycans.

Aspergillus nidulans is a well-established nonpathogenic laboratory model for the opportunistic mycopathogen, A. fumigatus. Some recent studies have focused on possible functional roles of glycosphingolipids (GSLs) in these fungi. It has been demonstrated that biosynthesis of glycosylinositol phosphorylceramides (GIPCs) is required for normal cell cycle progression and polarized growth in A. nidulans (Cheng, J., T.-S. Park, A. S. Fischl, and X. S. Ye. 2001. Mol. Cell Biol. 21: 6198-6209); however, the structures of A. nidulans GIPCs were not addressed in that study, nor were the functional significance of individual structural variants and the downstream steps in their biosynthesis. To initiate such studies, acidic GSL components (designated An-2, -3, and -5) were isolated from A. nidulans and subjected to structural characterization by a combination of one-dimensional (1-D) and 2-D NMR spectroscopy, electrospray ionization-mass spectrometry (ESI-MS), ESI-MS/collision-induced decomposition-MS (MS/CID-MS), ESI-pseudo-[CID-MS]2, and gas chromatography-MS methods. All three were determined to be GIPCs, with mannose as the only monosaccharide present in the headgroup glycans; An-2 and An-3 were identified as di- and trimannosyl inositol phosphorylceramides (IPCs) with the structures Man alpha 1-->3Man alpha 1-->2Ins1-P-1Cer and Man alpha 1-->3(Man alpha 1-->6)Man alpha 1-->2Ins1-P-1Cer, respectively (where Ins = myo-inositol, P = phosphodiester, and Cer = ceramide). An-5 was partially characterized, and is proposed to be a pentamannosyl IPC, based on the trimannosyl core structure of An-3.

Aspergillus nidulans↗
PubMed2003

The Aspergillus nidulans swoC1 mutant shows defects in growth and development.

Previous work identified swoC1 as a single-gene mutant with defects in polarity establishment. In this study swoC1 was shown to have defects in endocytosis, compartmentation, nuclear distribution, and conidiation. Temperature-shift experiments showed that the swoC1 mutant establishes multiple random sites of germ tube emergence. Surprisingly, these experiments also showed that even a slight delay in polarity establishment causes defects in later vegetative growth and asexual reproduction. The swoC gene was mapped to the centromere of chromosome III and cloned by complementation of the temperature-sensitive phenotype. The predicted SwoCp is homologous to rRNA pseudouridine synthases of yeast (Cbf5p) and humans (Dkc1p). However, neither rRNA pseudouridine synthesis nor rRNA processing appears to be affected in the swoC1 mutant. The swoC1 mutation occurs in the putative RNA-binding domain upstream of the C terminus, leaving the N-terminal TRUB catalytic domain intact. Interestingly, while deletion of the swoC gene was lethal in A. nidulans, the C terminus, including NLS, microtubule-binding, and coiled-coil domains, was dispensable for growth. SwoCp likely plays an important role in polar growth and nuclear distribution in A. nidulans, functions not yet described for its homologs.

Amino Acid Sequence↗
PubMed2003

SwoHp, a nucleoside diphosphate kinase, is essential in Aspergillus nidulans.

The temperature-sensitive swoH1 mutant of Aspergillus nidulans was previously identified in a screen for mutants with defects in polar growth. In the present work, we found that the swoH1 mutant swelled, lysed, and did not produce conidia during extended incubation at the restrictive temperature. When shifted from the permissive to the restrictive temperature, swoH1 showed the temperature-sensitive swelling phenotype only after 8 h at the higher temperature. The swoH gene was mapped to chromosome II and cloned by complementation of the temperature-sensitive phenotype. The sequence showed that swoH encodes a homologue of nucleoside diphosphate kinases (NDKs) from other organisms. Deletion experiments showed that the swoH gene is essential. A hemagglutinin-SwoHp fusion complemented the mutant phenotype, and the purified fusion protein possessed phosphate transferase activity in thin-layer chromatography assays. Sequencing of the mutant allele showed a predicted V83F change. Structural modeling suggested that the swoH1 mutation would lead to perturbation of the NDK active site. Crude cell extracts from the swoH1 mutant grown at the permissive temperature had approximately 20% of the NDK activity seen in the wild type and did not show any decrease in activity when assayed at higher temperatures. Though the data are not conclusive, the lack of temperature-sensitive NDK activity in the swoH1 mutant raises the intriguing possibility that the SwoH NDK is required for growth at elevated temperatures rather than for polarity maintenance.

Alleles↗
PubMed2002

Disruption of the glucosylceramide biosynthetic pathway in Aspergillus nidulans and Aspergillus fumigatus by inhibitors of UDP-Glc:ceramide glucosyltransferase strongly affects spore germination, cell cycle, and hyphal growth.

The opportunistic mycopathogen Aspergillus fumigatus expresses both glucosylceramide and galactosylceramide (GlcCer and GalCer), but their functional significance in Aspergillus species is unknown. We here identified and characterized a GlcCer from Aspergillus nidulans, a non-pathogenic model fungus. Involvement of GlcCer in fungal development was tested on both species using a family of compounds known to inhibit GlcCer synthase in mammals. Two analogs, D-threo-1-phenyl-2-palmitoyl-3-pyrrolidinopropanol (P4) and D-threo-3',4'-ethylenedioxy-P4, strongly inhibited germination and hyphal growth. Neutral lipids from A. fumigatus cultured in the presence of these inhibitors displayed a significantly reduced GlcCer/GalCer ratio. These results suggest that synthesis of GlcCer is essential for normal development of A. fumigatus and A. nidulans.

Antifungal Agents↗
PubMed2002

Aspergillus nidulans polarity mutant swoA is complemented by protein O-mannosyltransferase pmtA.

Previously swoA was identified in Aspergillus nidulans as a single locus, temperature-sensitive (ts) mutant aberrant in polarity maintenance. swoA was complemented by transformation with a plasmid genomic library. The sequence of the complementing gene was identical to a previously submitted GenBank sequence for pmtA, a protein O-mannosyltransferase. The pmtA/swoA-2 gene hybridized to three cosmids that are located on chromosome V and the swoA mutation was mitotically mapped to chromosome V. PMTs are endoplasmic reticulum-resident proteins responsible for the first step of O-glycosylation. Structural predictions suggest that PmtA contains seven membrane spans similar to PMTs from Saccharomyces cerevisiae and other organisms. Phylogenetic analysis indicates that PmtA is most closely related to the S. cerevisiae sub-family of PMTs containing Pmt2, Pmt3 and Pmt6. The mutant pmtA/swoA-2 locus contained a lesion that changed Y662 to a stop codon, eliminating the final membrane spanning domain and interrupting a domain essential for function in other PMTs.

Amino Acid Sequence↗
PubMed2002

Polarity in filamentous fungi: establishment, maintenance and new axes.

Germ tube emergence in filamentous fungi appears to be similar to bud emergence in yeast. Several key proteins (e.g. Cdc42, septins, Bni1 formin, Rho1 and Rho3) play common roles in polarity establishment and early polarity maintenance in both processes. Although germ tube extension, which can be thought of as extreme polarity maintenance, uses some of the same genes, they are likely to be regulated differently. Mutations in polarity maintenance genes often lead to a split tip in filamentous fungi, a phenotype without an analogue in yeast. Cell cycle regulation differs between tip splitting and subapical branching, but in both processes filamentous fungi maintain several axes of polar growth simultaneously.

Aspergillus nidulans↗
PubMed2002

Aspergillus nidulans septin AspB plays pre- and postmitotic roles in septum, branch, and conidiophore development.

Members of the septin family of proteins act as organizational scaffolds in areas of cell division and new growth in a variety of organisms. Herein, we show that in the filamentous fungus Aspergillus nidulans, the septin AspB is important for cellular division, branching, and conidiation both pre- and postmitotically. AspB localizes postmitotically to the septation site with an underlying polarity that is evident as cytokinesis progresses. This localization at the septation site is dependent on actin and occurs before the cross-wall is visible. AspB localizes premitotically as a ring at sites of branching and secondary germ tube emergence. It is the only known branch site marker. In addition, AspB is found at several stages during the development of the asexual reproductive structure, the conidiophore. It localizes transiently to the vesicle/metula and metula/phialide interfaces, and persistently to the phialide/conidiospore interface. A temperature-sensitive mutant of AspB shows phenotypic abnormalities, including irregular septa, high numbers of branches, and immature asexual reproductive structures.

Actins↗
PubMed2002

Aspergillus nidulans swoF encodes an N-myristoyl transferase.

Polar growth is a fundamental process in filamentous fungi and is necessary for disease initiation in many pathogenic systems. Previously, swoF was identified in Aspergillus nidulans as a single-locus, temperature-sensitive (ts) mutant aberrant in both polarity establishment and polarity maintenance. The swoF gene was cloned by complementation of the ts phenotype and sequenced. The derived protein sequence had high identity with N-myristoyl transferases (NMTs) found in fungi, plants, and animals. In addition, wild-type growth at restrictive temperature was partially restored by the addition of myristic acid to the growth medium. Sequencing revealed that the mutation in swoF changes the conserved aspartic acid 369 to a tyrosine. The predicted A. nidulans SwoF protein, SwoFp, was homology modeled based on crystal structures of NMTs from Saccharomyces cerevisiae and Candida albicans. The D369Y swoF mutation is on the opposite face of the protein, distal to the myristoyl coenzyme A and peptide substrate binding sites. In wild-type NMTs, D369 appears to stabilize a structural beta-strand bend through two hydrogen bonds and an ionic interaction. These stabilizing bonds are abolished in the D369Y mutant. We hypothesize that a substrate of SwoFp must be myristoylated for proper polarity establishment and maintenance. The mutation prevents the proper function of SwoFp at restrictive temperature and thus blocks polar growth.

Acyltransferases↗
PubMed2000

Landmarks in the early duplication cycles of Aspergillus fumigatus and Aspergillus nidulans: polarity, germ tube emergence and septation.

When the spores of filamentous fungi break dormancy, nuclear division is accompanied by a series of ordered morphological events including the switch from isotropic to polar growth, the emergence of a second germ tube from the conidium and septation. Correlation of these morphological events with nuclear number allows them to serve as duplication cycle landmarks. Early duplication cycle landmarks have been characterized in Aspergillus nidulans, but not in other filamentous fungi. To learn more about duplication cycle control in filamentous fungi, a study was undertaken to compare the timing of landmarks in Aspergillus fumigatus and A. nidulans. Nuclear duplication took approximately 45 min in A. fumigatus, with mitosis occupying roughly 5% of this period. Under the same conditions, nuclear duplication in A. nidulans took approximately 60 min, with mitosis occupying roughly 4% of this period. In A. fumigatus the isotropic to polar switch preceded the first mitosis in 22% of cells, while in A. nidulans the isotropic to polar switch did not occur until after the first mitosis. In both A. fumigatus and A. nidulans the earliest emergence of a second germ tube from the conidium occurred after the third mitotic division. However, by the fifth mitosis only 19% of A. fumigatus conidia had a second germ tube, compared to 98% of A. nidulans conidia. In both A. fumigatus and A. nidulans, formation of the first septum occurred after the fourth mitotic division. In all experiments a few cells lagged behind the others in nuclear number. In this delayed group, it was common to see landmark events at an earlier mitotic division. Differences in nuclear number when identical landmarks occur in A. fumigatus versus A. nidulans, and uncoupling of mitotic division and landmarks in delayed cells suggest that nuclear division and morphogenesis lie in parallel pathways, perhaps coordinated by checkpoints.

Aspergillus fumigatus↗