Publications by year
Bedekovic T, Brand AC
(2022). Microfabrication and its use in investigating fungal biology. Mol Microbiol
Microfabrication and its use in investigating fungal biology.
Advances in microfabrication technology, and its increasing accessibility, allow us to explore fungal biology as never before. By coupling molecular genetics with fluorescence live-cell imaging in custom-designed chambers, we can now probe single yeast cell responses to changing conditions over a lifetime, characterise population heterogeneity and investigate its underlying causes. By growing filamentous fungi in complex physical environments, we can identify cross-species commonalities, reveal species-specific growth responses and examine physiological differences relevant to diverse fungal lifestyles. As affordability and expertise broadens, microfluidic platforms will become a standard technique for examining the role of fungi in cross-kingdom interactions, ranging from rhizosphere to microbiome to interconnected human organ systems. This review brings together the perspectives already gained from studying fungal biology in microfabricated systems and outlines their potential in understanding the role of fungi in the environment, health and disease. Abstract
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da Silva Dantas A, Nogueira F, Lee KK, Walker LA, Edmondson M, Brand AC, Lenardon MD, Gow NAR
(2021). Crosstalk between the calcineurin and cell wall integrity pathways prevents chitin overexpression in Candida albicans. J Cell Sci
Crosstalk between the calcineurin and cell wall integrity pathways prevents chitin overexpression in Candida albicans.
Echinocandins such as caspofungin are frontline antifungal drugs that compromise β-1,3 glucan synthesis in the cell wall. Recent reports have shown that fungal cells can resist killing by caspofungin by upregulation of chitin synthesis, thereby sustaining cell wall integrity (CWI). When echinocandins are removed, the chitin content of cells quickly returns to basal levels, suggesting that there is a fitness cost associated with having elevated levels of chitin in the cell wall. We show here that simultaneous activation of the calcineurin and CWI pathways generates a subpopulation of Candida albicans yeast cells that have supra-normal chitin levels interspersed throughout the inner and outer cell wall, and that these cells are non-viable, perhaps due to loss of wall elasticity required for cell expansion and growth. Mutations in the Ca2+-calcineurin pathway prevented the formation of these non-viable supra-high chitin cells by negatively regulating chitin synthesis driven by the CWI pathway. The Ca2+-calcineurin pathway may therefore act as an attenuator that prevents the overproduction of chitin by coordinating both chitin upregulation and negative regulation of the CWI signaling pathway. This article has an associated First Person interview with the first author of the paper. Abstract
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Wehmeier S, Morrison E, Plato A, Raab A, Feldmann J, Bedekovic T, Wilson D, Brand AC
(2020). Multi trace element profiling in pathogenic and non-pathogenic fungi. Fungal Biol
Multi trace element profiling in pathogenic and non-pathogenic fungi.
Maintaining appropriate levels of trace elements during infection of a host is essential for microbial pathogenicity. Here we compared the uptake of 10 trace elements from 3 commonly-used laboratory media by 3 pathogens, Candida albicans, Cryptococcus neoformans and Aspergillus fumigatus, and a model yeast, Saccharomyces cerevisiae. The trace element composition of the yeasts, C. albicans, C. neoformans and S. cerevisiae, grown in rich (YPD) medium, differed primarily in P, S, Fe, Zn and Co. Speciation analysis of the intracellular fraction, which indicates the size of the organic ligands with which trace elements are complexed, showed that the ligands for S were similar in the three fungi but there were significant differences in binding partners for Fe and Zn between C. neoformans and S.cerevisiae. The profile for Cu varied across the 3 yeast species. In a comparison of C. albicans and A. fumigatus hyphae, the former showed higher Fe, Cu, Zn and Mn, while A. fumigatus contained higher P, S Ca and Mo. Washing C. albicans cells with the cell-impermeable chelator, EGTA, depleted 50-90 % of cellular Ca, suggesting that a large proportion of this cation is stored in the cell wall. Treatment with the cell wall stressor, Calcofluor White (CFW), alone had little effect on the elemental profile whilst combined Ca + CFW stress resulted in high cellular Cu and very high Ca. Together our data enhance our understanding of trace element uptake by pathogenic fungi and provide evidence for the cell wall as an important storage organelle for Ca. Abstract
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Bedekovic T, Agnew E, Brand AC
(2020). Rsr1 Palmitoylation and GTPase Activity Status Differentially Coordinate Nuclear, Septin, and Vacuole Dynamics in Candida albicans. mBio
Rsr1 Palmitoylation and GTPase Activity Status Differentially Coordinate Nuclear, Septin, and Vacuole Dynamics in Candida albicans
. Understanding how single eukaryotic cells self-organize to replicate and migrate is relevant to health and disease. In the fungal pathogen,
. Candida albicans
. the small GTPase, Rsr1, guides the directional growth of hyphae that invade human tissue during life-threatening infections. Rsr1 is a Ras-like GTPase and a homolog of the conserved Rap1 subfamily, which directs migration in mammalian cells. Research into how this single GTPase delivers complex intracellular patterning is challenging established views of GTPase regulation, trafficking, and interaction. Here, we show that Rsr1 directly and indirectly coordinates the spatial and temporal development of key intracellular macrostructures, including septum formation and closure, vacuole dynamics, and nuclear division and segregation, as well as whole-cell morphology by determining branching patterns. Furthermore, we categorize these functions by differential Rsr1 localization and activity state and provide evidence to support the emerging view that the cytosolic pool of Ras-like GTPases is functionally active.
Brand A, Bedekovic T, Agnew E
(2020). Rsr1 palmitoylation and GTPase activity-status differentially co-ordinate nuclear, septin and vacuole dynamics in Candida albicans.
Rsr1 palmitoylation and GTPase activity-status differentially co-ordinate nuclear, septin and vacuole dynamics in Candida albicans
Directional growth and tissue invasion by hyphae of the pathogenic fungus, Candida albicans, are disrupted by deletion of the small GTPase, Rsr1, which localises Cdc42 and its kinase, Cla4, to the site of polarized growth. We investigated additional abnormalities observed in rsr1Δ hyphae, including vacuole development, cytoplasm inheritance, mitochondrial morphology, septin ring organization, nuclear division and migration, and branching-frequency, which together demonstrate a fundamental role for Rsr1 in cellular organization. Rsr1 contains a C-terminal CCAAX-box, which putatively undergoes both reversible palmitoylation, and farnesylation for entry into the secretory pathway. We expressed variants of Rsr1 with mutated C244 or C245, or which lacked GTPase activity (Rsr1K16N and Rsr1G12V) in the rsr1Δ background and compared the resulting phenotypes with mutants lacking Bud5 (Rsr1GEF), Bud2 (Rsr1GAP) or Cla4. Bud5 was required only for cell size and bud-site selection in yeast, suggesting there are alternative activators for Rsr1 in hyphae. Septin ring and vacuole dynamics were restored by expression of unpalmitoylated Rsr1C244S, which localized to endomembranes, but not by cytoplasmic Rsr1C245A or GTP/GDP-locked Rsr1, suggesting Rsr1 functions at intracellular membranes in addition to the plasma-membrane. Rsr1K16N or cytoplasmic Rsr1C245A restored normal nuclear division, but not septin ring or vacuole dynamics. Rsr1-GDP therefore plays a specific role in suppressing Start, which can be signalled from the cytosol. Via differential palmitoylation and activity states, Rsr1 operates at diverse cell sites to orchestrate proper nuclear division and inheritance during constitutive polarized growth. As cla4Δ phenocopied rsr1Δ, it is likely these functions involve Cdc42-Cla4 activity. Abstract
Alder-Rangel A, Idnurm A, Brand AC, Brown AJP, Gorbushina A, Kelliher CM, Campos CB, Levin DE, Bell-Pedersen D, Dadachova E, et al (2020). The Third International Symposium on Fungal Stress – ISFUS. Fungal Biology, 124(5), 235-252.
Martin-Yken H, Bedekovic T, Brand AC, Richard ML, Znaidi S, d'Enfert C, Dague E (2018). A conserved fungal hub protein involved in adhesion and drug resistance in the human pathogen Candida albicans. The Cell Surface, 4, 10-19.
du Pré S, Beckmann N, Almeida MC, Sibley GEM, Law D, Brand AC, Birch M, Read ND, Oliver JD
(2018). Effect of the Novel Antifungal Drug F901318 (Olorofim) on Growth and Viability of Aspergillus fumigatus. Antimicrobial Agents and Chemotherapy
Effect of the Novel Antifungal Drug F901318 (Olorofim) on Growth and Viability of Aspergillus fumigatus
. F901318 (olorofim) is a novel antifungal drug that is highly active against
. species. Belonging to a new class of antifungals called the orotomides, F901318 targets dihydroorotate dehydrogenase (DHODH) in the
. de novo
. pyrimidine biosynthesis pathway.
Garcia AML, Almeida MC, Bedekovic T, Brand A
(2018). The Cst20 and Ptk2 kinases are involved in hyphal steering and thigmotropic responses in Candida albicans. Author URL
Riquelme M, Aime MC, Branco S, Brand A, Brown A, Glass NL, Kahmann R, Momany M, Rokas A, Trail F, et al (2018). The power of discussion: Support for women at the fungal Gordon Research Conference. Fungal Genetics and Biology, 121, 65-67.
Almeida MC, Brand AC
(2017). Thigmo Responses: the Fungal Sense of Touch. In (Ed)
, American Society for Microbiology.
Thigmo Responses: the Fungal Sense of Touch
Almeida MC, Brand AC
(2017). Thigmo responses: the fungal sense of touch. In (Ed) The Fungal Kingdom
Thigmo responses: the fungal sense of touch
Thomson DD, Berman J, Brand AC (2016). High frame-rate resolution of cell division during Candida albicans filamentation. Fungal Genetics and Biology, 88, 54-58.
Chavez-Dozal AA, Bernardo SM, Rane HS, Herrera G, Kulkarny V, Wagener J, Cunningham I, Brand AC, Gow NAR, Lee SA, et al (2015). Retraction for Chavez-Dozal et al. the Candida albicans Exocyst Subunit Sec6 Contributes to Cell Wall Integrity and is a Determinant of Hyphal Branching. Eukaryotic Cell, 14(12).
Chavez-Dozal AA, Bernardo SM, Rane HS, Herrera G, Kulkarny V, Wagener J, Cunningham I, Brand AC, Gow NAR, Leea SA, et al
(2015). The Candida albicans exocyst subunit Sec6 contributes to cell wall integrity and is a determinant of hyphal branching. Eukaryotic Cell
The Candida albicans exocyst subunit Sec6 contributes to cell wall integrity and is a determinant of hyphal branching
The yeast exocyst is a multiprotein complex comprised of eight subunits (Sec3, Sec5, Sec6, Sec8, Sec10, Sec15, Exo70, and Exo84) which orchestrates trafficking of exocytic vesicles to specific docking sites on the plasma membrane during polarized secretion. To study SEC6 function in Candida albicans, we generated a conditional mutant strain in which SEC6 was placed under the control of a tetracycline-regulated promoter. In the repressed state, the tetR-SEC6 mutant strain (denoted tSEC6) was viable for up to 27 h; thus, all phenotypic analyses were performed at 24 h or earlier. Strain tSEC6 under repressing conditions had readily apparent defects in cytokinesis and endocytosis and accumulated both post-Golgi apparatus secretory vesicles and structures suggestive of late endosomes. Strain tSEC6 was markedly defective in secretion of aspartyl proteases and lipases as well as filamentation under repressing conditions. Lack of SEC6 expression resulted in markedly reduced lateral hyphal branching, which requires the establishment of a new axis of polarized secretion. Aberrant localization of chitin at the septum and increased resistance to zymolyase activity were observed, suggesting that C. albicans Sec6 plays an important role in mediating trafficking and delivery of cell wall components. The tSEC6 mutant was also markedly defective in macrophage killing, indicating a role of SEC6 in C. albicans virulence. Taken together, these studies indicate that the late secretory protein Sec6 is required for polarized secretion, hyphal morphogenesis, and the pathogenesis of C. albicans. Abstract
Brand AC, Morrison E, Milne S, Gonia S, Gale CA, Gow NAR
(2014). Cdc42 GTPase dynamics control directional growth responses. Proceedings of the National Academy of Sciences of the United States of America
Cdc42 GTPase dynamics control directional growth responses
Polarized cells reorient their direction of growth in response toenvironmental cues. In the fungus Candida albicans, the Rho-familysmall GTPase, Cdc42, is essential for polarized hyphal growth andCa2+ influx is required for the tropic responses of hyphae to environmental cues, but the regulatory link between these systems isunclear. In this study, the interaction between Ca2+ influx and Cdc42polarity-complex dynamics was investigated using hyphal galvano-tropic and thigmotropic responses as reporter systems. During polarity establishment in an applied electric field, cathodal emergenceof hyphae was lost when either of the two Cdc42 apical recyclingpathways was disrupted by deletion of Rdi1, a guanine nucleotidedissociation inhibitor, or Bnr1, a formin, but was completely restored by extracellular Ca2+. Loss of the Cdc42 GTPase activatingproteins, Rga2 and Bem3, also abolished cathodal polarization, butthis was not rescued by Ca2+. Expression of GTP-locked Cdc42 reversed the polarity of hypha emergence from cathodal to anodal, aneffect augmented by Ca2+. The cathodal directional cue thereforerequires Cdc42 GTP hydrolysis. Ca2+ influx amplifies Cdc42-mediateddirectional growth signals, in part by augmenting Cdc42 apical trafficking. The Ca2+-binding EF-hand motif in Cdc24, the Cdc42 activator, was essential for growth in yeast cells but not in establishedhyphae. The Cdc24 EF-hand motif is therefore essential for polarityestablishment but not for polarity maintenance. Abstract
Thomson DD, Wehmeier S, Byfield FJ, Janmey PA, Caballero-Lima D, Crossley A, Brand AC (2014). Contact-induced apical asymmetry drives the thigmotropic responses of<i>C</i><i>andida albicans</i>hyphae. Cellular Microbiology, 17(3), 342-354.
Gonia S, Norton J, Watanaskul L, Pulver R, Morrison E, Brand A, Gale CA (2013). Rax2 is important for directional establishment of growth sites, but not for reorientation of growth axes, during Candida albicans hyphal morphogenesis. Fungal Genetics and Biology, 56, 116-124.
Ene IV, Adya AK, Wehmeier S, Brand AC, Maccallum DM, Gow NAR, Brown AJP
(2012). Host carbon sources modulate cell wall architecture, drug resistance and virulence in a fungal pathogen. Cellular Microbiology
Host carbon sources modulate cell wall architecture, drug resistance and virulence in a fungal pathogen
The survival of all microbes depends upon their ability to respond to environmental challenges. To establish infection, pathogens such as Candida albicans must mount effective stress responses to counter host defences while adapting to dynamic changes in nutrient status within host niches. Studies of C.albicans stress adaptation have generally been performed on glucose-grown cells, leaving the effects of alternative carbon sources upon stress resistance largely unexplored. We have shown that growth on alternative carbon sources, such as lactate, strongly influence the resistance of C.albicans to antifungal drugs, osmotic and cell wall stresses. Similar trends were observed in clinical isolates and other pathogenic Candida species. The increased stress resistance of C.albicans was not dependent on key stress (Hog1) and cell integrity (Mkc1) signalling pathways. Instead, increased stress resistance was promoted by major changes in the architecture and biophysical properties of the cell wall. Glucose- and lactate-grown cells displayed significant differences in cell wall mass, ultrastructure, elasticity and adhesion. Changes in carbon source also altered the virulence of C.albicans in models of systemic candidiasis and vaginitis, confirming the importance of alternative carbon sources within host niches during C.albicans infections. The major fungal pathogen of humans, Candida albicans, often exploits alternative carbon sources to grow and colonise glucose-poor niches in its host. Most investigations of stress adaptation have examined glucose-grown cells, but we demonstrate that growth on physiologically relevant carbon sources such as lactate alters C. albicans cell wall architecture and that this affects the resistance of cells to stress and antifungal drugs. Furthermore, growth on alternative carbon sources alters the pathogenicity of C. albicans during systemic and mucosal infections. © 2012 Blackwell Publishing Ltd. Abstract
(2012). Hyphal growth in human fungal pathogens and its role in virulence. International Journal of Microbiology
Hyphal growth in human fungal pathogens and its role in virulence
Most of the fungal species that infect humans can grow in more than one morphological form but only a subset of pathogens produce filamentous hyphae during the infection process. This subset is phylogenetically unrelated and includes the commonly carried yeasts, Candida albicans, C. dubliniensis, and Malassezia spp. and the acquired pathogens, Aspergillus fumigatus and dermatophytes such as Trichophyton rubrum and T. mentagrophytes. The primary function of hypha formation in these opportunistic pathogens is to invade the substrate they are adhered to, whether biotic or abiotic, but other functions include the directional translocation between host environments, consolidation of the colony, nutrient acquisition and the formation of 3-dimensional matrices. To support these functions, polarised hyphal growth is co-regulated with other factors that are essential for normal hypha function in vivo. Abstract
Brand AC, Maccallum DM (2012). Preface.
Brand A, Gow NAR
(2012). Tropic orientation responses of pathogenic fungi.
Tropic orientation responses of pathogenic fungi
Chen Y-L, Brand A, Morrison EL, Silao FGS, Bigol UG, Malbas FF, Nett JE, Andes DR, Solis NV, Filler SG, et al
(2011). Calcineurin controls drug tolerance, hyphal growth, and virulence in Candida dubliniensis. Eukaryot Cell
Calcineurin controls drug tolerance, hyphal growth, and virulence in Candida dubliniensis.
Candida dubliniensis is an emerging pathogenic yeast species closely related to Candida albicans and frequently found colonizing or infecting the oral cavities of HIV/AIDS patients. Drug resistance during C. dubliniensis infection is common and constitutes a significant therapeutic challenge. The calcineurin inhibitor FK506 exhibits synergistic fungicidal activity with azoles or echinocandins in the fungal pathogens C. albicans, Cryptococcus neoformans, and Aspergillus fumigatus. In this study, we show that calcineurin is required for cell wall integrity and wild-type tolerance of C. dubliniensis to azoles and echinocandins; hence, these drugs are candidates for combination therapy with calcineurin inhibitors. In contrast to C. albicans, in which the roles of calcineurin and Crz1 in hyphal growth are unclear, here we show that calcineurin and Crz1 play a clearly demonstrable role in hyphal growth in response to nutrient limitation in C. dubliniensis. We further demonstrate that thigmotropism is controlled by Crz1, but not calcineurin, in C. dubliniensis. Similar to C. albicans, C. dubliniensis calcineurin enhances survival in serum. C. dubliniensis calcineurin and crz1/crz1 mutants exhibit attenuated virulence in a murine systemic infection model, likely attributable to defects in cell wall integrity, hyphal growth, and serum survival. Furthermore, we show that C. dubliniensis calcineurin mutants are unable to establish murine ocular infection or form biofilms in a rat denture model. That calcineurin is required for drug tolerance and virulence makes fungus-specific calcineurin inhibitors attractive candidates for combination therapy with azoles or echinocandins against emerging C. dubliniensis infections. Abstract
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Yang M, Brand A, Srikantha T, Daniels KJ, Soll DR, Gow NAR
(2011). Fig1 facilitates calcium influx and localizes to membranes destined to undergo fusion during mating in Candida albicans. Eukaryot Cell
Fig1 facilitates calcium influx and localizes to membranes destined to undergo fusion during mating in Candida albicans.
Few mating-regulated genes have been characterized in Candida albicans. C. albicans FIG1 (CaFIG1) is a fungus-specific and mating-induced gene encoding a putative 4-transmembrane domain protein that shares sequence similarities with members of the claudin superfamily. In Saccharomyces cerevisiae, Fig1 is required for shmoo fusion and is upregulated in response to mating pheromones. Expression of CaFIG1 was also strongly activated in the presence of cells of the opposite mating type. CaFig1-green fluorescent protein (GFP) was visible only during the mating response, when it localized predominantly to the plasma membrane and perinuclear zone in mating projections and daughter cells. At the plasma membrane, CaFig1-GFP was visualized as discontinuous zones, but the distribution of perinuclear CaFig1-GFP was homogeneous. Exposure to pheromone induced a 5-fold increase in Ca(2+) uptake in mating-competent opaque cells. Uptake was reduced substantially in the fig1Δ null mutant. CaFig1 is therefore involved in Ca(2+) influx and localizes to membranes that are destined to undergo fusion during mating. Abstract
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Brand A (2011). Host Fungus Interactions Vol 1 & 2.
Mora-Montes HM, Bates S, Netea MG, Castillo L, Brand A, Buurman ET, Díaz-Jiménez DF, Jan Kullberg B, Brown AJP, Odds FC, et al
(2010). A multifunctional mannosyltransferase family in Candida albicans determines cell wall mannan structure and host-fungus interactions. J Biol Chem
A multifunctional mannosyltransferase family in Candida albicans determines cell wall mannan structure and host-fungus interactions.
The cell wall proteins of fungi are modified by N- and O-linked mannosylation and phosphomannosylation, resulting in changes to the physical and immunological properties of the cell. Glycosylation of cell wall proteins involves the activities of families of endoplasmic reticulum and Golgi-located glycosyl transferases whose activities are difficult to infer through bioinformatics. The Candida albicans MNT1/KRE2 mannosyl transferase family is represented by five members. We showed previously that Mnt1 and Mnt2 are involved in O-linked mannosylation and are required for virulence. Here, the role of C. albicans MNT3, MNT4, and MNT5 was determined by generating single and multiple MnTDelta null mutants and by functional complementation experiments in Saccharomyces cerevisiae. CaMnt3, CaMnt4, and CaMnt5 did not participate in O-linked mannosylation, but CaMnt3 and CaMnt5 had redundant activities in phosphomannosylation and were responsible for attachment of approximately half of the phosphomannan attached to N-linked mannans. CaMnt4 and CaMnt5 participated in N-mannan branching. Deletion of CaMNT3, CaMNT4, and CaMNT5 affected the growth rate and virulence of C. albicans, affected the recognition of the yeast by human monocytes and cytokine stimulation, and led to increased cell wall chitin content and exposure of beta-glucan at the cell wall surface. Therefore, the MNT1/KRE2 gene family participates in three types of protein mannosylation in C. albicans, and these modifications play vital roles in fungal cell wall structure and cell surface recognition by the innate immune system. Abstract
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Brand A, Lee K, Veses V, Gow NAR (2009). Calcium homeostasis is required for contact-dependent helical and sinusoidal tip growth in. Candida albicans. hyphae. Molecular Microbiology
Brand A, Lee K, Veses V, Gow NAR
(2009). Calcium homeostasis is required for contact-dependent helical and sinusoidal tip growth in Candida albicans hyphae. Molecular Microbiology
Calcium homeostasis is required for contact-dependent helical and sinusoidal tip growth in Candida albicans hyphae
Hyphae of the dimorphic fungus, Candida albicans, exhibit directional tip responses when grown in contact with surfaces. On hard surfaces or in liquid media, the trajectory of hyphal growth is typically linear, with tip re-orientation events limited to encounters with topographical features (thigmotropism). In contrast, when grown on semisolid surfaces, the tips of C. albicans hyphae grow in an oscillatory manner to form regular two-dimensional sinusoidal curves and three-dimensional helices. We show that, like thigmotropism, initiation of directional tip oscillation in C. albicans hyphae is severely attenuated when Ca2+ homeostasis is perturbed. Chelation of extracellular Ca2+ or deletion of the Ca2+ transporters that modulate cytosolic [Ca2+] (Mid1, Cch1 or Pmr1) did not affect hyphal length but curve formation was severely reduced in mid1Δ and cch1Δ and abolished in pmr1Δ. Sinusoidal hypha morphology was altered in the mid1Δ, chs3Δ and heterozygous pmr1Δ/PMR1 strains. Treatments that affect cell wall integrity, changes in surface mannosylation or the provision of additional carbon sources had significant but less pronounced effects on oscillatory growth. The induction of two- and three-dimensional sinusoidal growth in wild-type C. albicans hyphae is therefore the consequence of mechanisms that involve Ca2+ influx and signalling rather than gross changes in the cell wall architecture. © 2009 Blackwell Publishing Ltd. Abstract
Brand A, Gow NA
(2009). Mechanisms of hypha orientation of fungi. Current Opinion in Microbiology
Mechanisms of hypha orientation of fungi
Hypha orientation is an essential aspect of polarised growth and the morphogenesis, spatial ecology and pathogenesis of fungi. The ability to re-orient tip growth in response to environmental cues is critical for colony ramification, the penetration of diverse host tissues and the formation of mating structures. Recent studies have begun to describe the molecular machinery regulating hypha orientation. Calcium signalling, the polarisome Bud1-GTPase module and the Tea cell-end marker proteins of the microtubule cytoskeleton, along with specific kinesins and sterol-rich apical microdomains, are involved in hypha orientation. Mutations that affect these processes generate normal-shaped, growing hyphae that have either abnormal meandering trajectories or attenuated tropic responses. Hyphal tip orientation and tip extension are, therefore, distinct regulatory mechanisms that operate in parallel during filamentous growth, thereby allowing fungi to orchestrate their reproduction in relation to gradients of effectors in their environments. © 2009 Elsevier Ltd. All rights reserved. Abstract
Brand A, Vacharaksa A, Bendel C, Norton J, Haynes P, Henry-Stanley M, Wells C, Ross K, Gow NAR, Gale CA, et al
(2008). An internal polarity landmark is important for externally induced hyphal behaviors in Candida albicans. Eukaryotic Cell
An internal polarity landmark is important for externally induced hyphal behaviors in Candida albicans
Directional growth is a function of polarized cells such as neurites, pollen tubes, and fungal hyphae. Correct orientation of the extending cell tip depends on signaling pathways and effectors that mediate asymmetric responses to specific environmental cues. In the hyphal form of the eukaryotic fungal pathogen Candida albicans, these responses include thigmotropism and galvanotropism (hyphal turning in response to changes in substrate topography and imposed electrical fields, respectively) and penetration into semisolid substrates. During vegetative growth in C. albicans, as in the model yeast Saccharomyces cerevisiae, the Ras-like GTPase Rsr1 mediates internal cellular cues to position new buds in a prespecified pattern on the mother cell cortex. Here, we demonstrate that Rsr1 is also important for hyphal tip orientation in response to the external environmental cues that induce thigmotropic and galvanotropic growth. In addition, Rsr1 is involved in hyphal interactions with epithelial cells in vitro and its deletion diminishes the hyphal invasion of kidney tissue during systemic infection. Thus, Rsr1, an internal polarity landmark in yeast, is also involved in polarized growth responses to asymmetric environmental signals, a paradigm that is different from that described for the homologous protein in S. cerevisiae. Rsr1 may thereby contribute to the pathogenesis of C. albicans infections by influencing hyphal tip responses triggered by interaction with host tissues. Copyright © 2008, American Society for Microbiology. All Rights Reserved. Abstract
Brand A, Barnes JD, Mackenzie KS, Odds FC, Gow NAR
(2008). Cell wall glycans and soluble factors determine the interactions between the hyphae of Candida albicans and Pseudomonas aeruginosa. FEMS Microbiology Letters
Cell wall glycans and soluble factors determine the interactions between the hyphae of Candida albicans and Pseudomonas aeruginosa
The fungus, Candida albicans, and the bacterium, Pseudomonas aeruginosa, are opportunistic human pathogens that have been coisolated from diverse body sites. Pseudomonas aeruginosa suppresses C. albicans proliferation in vitro and potentially in vivo but it is the C. albicans hyphae that are killed while yeast cells are not. We show that hyphal killing involves both contact-mediated and soluble factors. Bacterial culture filtrates contained heat-labile soluble factors that killed C. albicans hyphae. In cocultures, localized points of hyphal lysis were observed, suggesting that adhesion and subsequent bacteria-mediated cell wall lysis is involved in the killing of C. albicans hyphae. The glycosylation status of the C. albicans cell wall affected the rate of contact-dependent killing because mutants with severely truncated O-linked, but not N-linked, glycans were hypersensitive to Pseudomonas-mediated killing. Deletion of HWP1, ALS3 or HYR1, which encode major hypha-associated cell wall proteins, had no effect on fungal susceptibility. © 2008 the Authors. Abstract
Brand A, Shanks S, Duncan VMS, Yang M, Mackenzie K, Gow NAR
(2007). Hyphal Orientation of Candida albicans is Regulated by a Calcium-Dependent Mechanism. Current Biology
Hyphal Orientation of Candida albicans is Regulated by a Calcium-Dependent Mechanism
Eukaryotic cells from fungal hyphae to neurites that grow by polarized extension must coordinate cell growth and cell orientation to enable them to exhibit growth tropisms and to respond to relevant environmental cues. Such cells generally maintain a tip-high Ca2+ cytoplasmic gradient, which is correlated with their ability to exhibit polarized tip growth and to respond to growth-directing extracellular signals [1-5]. In yeast and other fungi, the polarisome, exocyst, Arp2/3, and Spitzenkörper protein complexes collectively orchestrate tip growth and cell polarity, but it is not clear whether these molecular complexes also regulate cell orientation or whether they are influenced by cytoplasmic Ca2+ gradients. Hyphae of the human pathogenic fungus Candida albicans reorient their growth axis in response to underlying surface topography (thigmotropism)  and imposed electric fields (galvanotropism) . The establishment and maintenance of directional growth in relation to these environmental cues was Ca2+ dependent. Tropisms were attenuated in media containing low Ca2+, or calcium-channel blockers, and in mutants where calcium channels or elements of the calcium signaling pathway were deleted. Therefore galvanotropism and thigmotropism may both be mediated by localized Ca2+ influx at sites of polarized growth via Ca2+ channels that are activated by appropriate environmental signals. © 2007 Elsevier Ltd. All rights reserved. Abstract
Munro CA, Bates S, Buurman ET, Hughes HB, MacCallum DM, Bertram G, Atrih A, Ferguson MAJ, Bain JM, Brand A, et al (2005). Mnt1p and Mnt2p of Candida albicans Are Partially Redundant α-1,2-Mannosyltransferases That Participate in O-Linked Mannosylation and Are Required for Adhesion and Virulence. Journal of Biological Chemistry, 280(2), 1051-1060.
Brand A, MacCallum DM, Brown AJP, Gow NAR, Odds FC
(2004). Ectopic expression of URA3 can influence the virulence phenotypes and proteome of Candida albicans but can be overcome by targeted reintegration of URA3 at the RPS10 locus. Eukaryotic Cell
Ectopic expression of URA3 can influence the virulence phenotypes and proteome of Candida albicans but can be overcome by targeted reintegration of URA3 at the RPS10 locus
Uridine auxotrophy, based on disruption of both URA3 alleles in diploid Candida albicans strain SC5314, has been widely used to select gene deletion mutants created in this fungus by "Ura-blasting" and PCR-mediated disruption. We compared wild-type URA3 expression with levels in mutant strains where URA3 was positioned either within deleted genes or at the highly expressed RPS10 locus. URA3 expression levels differed significantly and correlated with the specific activity of Ura3p, orotidine 5′-monophosphate decarboxylase. Reduced URA3 expression following integration at the GCN4 locus was associated with an attenuation of virulence. Furthermore, a comparison of the SC5314 (URA3) and CAI-4 (ura3) proteomes revealed that inactivation of URA3 caused significant changes in the levels of 14 other proteins. The protein levels of all except one were partially or fully restored by the reintegration of a single copy of URA3 at the RPS10 locus. Transcript levels of genes expressed ectopically at this locus in reconstituted heterozygous mutants also matched the levels found when the genes were expressed at their native loci. Therefore, phenotypic changes in C. albicans can be associated with the selectable marker rather than the target gene. Reintegration of URA3 at an appropriate expression locus such as RPS10 can offset most problems related to the phenotypic changes associated with gene knockout methodologies. Abstract