Cytosolic free calcium dynamics as related to hyphal and colony growth in the filamentous fungal pathogen Colletotrichum graminicola

Lange, Mario; Peiter, Edgar

doi:10.1016/j.fgb.2016.04.001

Cited by 8 publications

(17 citation statements)

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“…As the CgTRPF proteins have similarities to the Ca 2+ -permeable TRPY1, which is known to regulate [Ca 2+ ] cyt homeostasis in yeast, we tested if they play a role for growth under Ca 2+ -limited conditions. To this end, the wild type and deletion strains were cultivated on Ca 2+ -depleted SC agar medium, containing 1.7 μM total Ca 2+ , and on Ca 2+ -replete SC medium supplemented with 900 μM Ca 2+ [ 45 ]. As previously reported [ 45 ], the wild type showed a growth reduction by around 20% on low-Ca 2+ medium ( Fig 7 ).…”

Section: Resultsmentioning

confidence: 99%

“…To induce the production of falcate conidia, the fungus was grown on oat meal agar (OMA) [ 45 ]. Colony growth assays were performed as described before [ 45 ]. Colony diameters were recorded daily for 4 to 10 days.…”

Section: Methodsmentioning

confidence: 99%

“…Sodium malate and sodium pectate were obtained by neutralizing DL-malic acid and pectic acid to pH 7.0 with NaOH, respectively. Ca 2+ -depleted SC medium was prepared as described before [ 45 ].…”

Section: Methodsmentioning

confidence: 99%

“…To test [Ca 2+ ] cyt responses of C . graminicola wild type and Δ Cgtrpf deletion strains to osmotic upshock, the strains were transformed with the pAEQS1-G418 plasmid [ 45 ] which encodes a codon-optimized Apoaequorin [ 56 ]. Transformants were selected by using G418 (geneticin; 600 μg mL -1 during transformation, 150 μg mL -1 during selection and single-spore isolation).…”

Section: Methodsmentioning

confidence: 99%

“…For [Ca 2+ ] cyt analyses in individual hyphae, wild type and Δ Cgtrpf deletion strains were transformed with a pGEM-T-PtrpC-nptII-TtrpC-PtoxB-YC3.6-TtrpC plasmid [ 45 ], encoding the FRET-based ratiometric Ca 2+ reporter protein Yellow Cameleon 3.6 (YC3.6) [ 57 ]. Measurements of tip-focussed [Ca 2+ ] cyt spikes during hyphal growth were performed as described before [ 45 ].…”

Section: Methodsmentioning

confidence: 99%

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The Transient Receptor Potential (TRP) Channel Family in Colletotrichum graminicola: A Molecular and Physiological Analysis

et al. 2016

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Calcium (Ca2+) is a universal second messenger in all higher organisms and centrally involved in the launch of responses to environmental stimuli. Ca2+ signals in the cytosol are initiated by the activation of Ca2+ channels in the plasma membrane and/or in endomembranes. Yeast (Saccharomyces cerevisiae) contains a Ca2+-permeable channel of the TRP family, TRPY1, which is localized in the vacuolar membrane and contributes to cytosolic free Ca2+ ([Ca2+]cyt) elevations, for example in response to osmotic upshock. A TRPY1 homologue in the rice blast fungus is known to be important for growth and pathogenicity. To determine the role of the TRP channel family in the maize pathogen Colletotrichum graminicola, proteins homologous to TRPY1 were searched. This identified not one, but four genes in the C. graminicola genome, which had putative orthologs in other fungi, and which we named CgTRPF1 through 4. The topology of the CgTRPF proteins resembled that of TRPY1, albeit with a variable number of transmembrane (TM) domains additional to the six-TM-domain core and a diverse arrangement of putatively Ca2+-binding acidic motifs. All CgTRPF genes were expressed in axenic culture and throughout the infection of maize. Like TRPY1, all TRPF proteins of C. graminicola were localized intracellularly, albeit three of them were found not in large vacuoles, but co-localized in vesicular structures. Deletion strains for the CgTRPF genes were not altered in processes thought to involve Ca2+ release from internal stores, i.e. spore germination, the utilization of complex carbon sources, and the generation of tip-focussed [Ca2+]cyt spikes. Heterologous expression of CgTRPF1 through 4 in a tryp1Δ yeast mutant revealed that none of the channels mediated the release of Ca2+ in response to osmotic upshock. Accordingly, aequorin-based [Ca2+]cyt measurements of C. graminicola showed that in this fungus, osmotic upshock-triggered [Ca2+]cyt elevations were generated entirely by influx of Ca2+ from the extracellular space. Cgtrpf mutants did not show pathogenicity defects in leaf infection assays. In summary, our study reveals major differences between different fungi in the contribution of TRP channels to Ca2+-mediated signal transduction.

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Section: Resultsmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

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The Transient Receptor Potential (TRP) Channel Family in Colletotrichum graminicola: A Molecular and Physiological Analysis

et al. 2016

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Calcium dynamics in tomato pollen tubes using the Yellow Cameleon 3.6 sensor

et al. 2017

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In vitro tomato pollen tubes show a cytoplasmic calcium gradient that oscillates with the same period as growth. Pollen tube growth requires coordination between the tip-focused cytoplasmic calcium concentration ([Ca]) gradient and the actin cytoskeleton. This [Ca] gradient is necessary for exocytosis of small vesicles, which contributes to the delivery of new membrane and cell wall at the pollen tube tip. The mechanisms that generate and maintain this [Ca] gradient are not completely understood. Here, we studied calcium dynamics in tomato (Solanum lycopersicum) pollen tubes using transgenic tomato plants expressing the Yellow Cameleon 3.6 gene under the pollen-specific promoter LAT52. We use tomato as an experimental model because tomato is a Solanaceous plant that is easy to transform, and has an excellent genomic database and genetic stock center, and unlike Arabidopsis, tomato pollen is a good system to do biochemistry. We found that tomato pollen tubes showed an oscillating tip-focused [Ca] gradient with the same period as growth. Then, we used a pharmacological approach to disturb the intracellular Ca homeostasis, evaluating how the [Ca] gradient, pollen germination and in vitro pollen tube growth were affected. We found that cyclopiazonic acid (CPA), a drug that inhibits plant P-type Ca-ATPases, increased [Ca] in the subapical zone, leading to the disappearance of the Ca oscillations and inhibition of pollen tube growth. In contrast, 2-aminoethoxydiphenyl borate (2-APB), an inhibitor of Ca released from the endoplasmic reticulum to the cytoplasm in animals cells, completely reduced [Ca] at the tip of the tube, blocked the gradient and arrested pollen tube growth. Although both drugs have antagonistic effects on [Ca], both inhibited pollen tube growth triggering the disappearance of the [Ca] gradient. When CPA and 2-APB were combined, their individual inhibitory effects on pollen tube growth were partially compensated. Finally, we found that GsMTx-4, a peptide from spider venom that blocks stretch-activated Ca channels, inhibited tomato pollen germination and had a heterogeneous effect on pollen tube growth, suggesting that these channels are also involved in the maintenance of the [Ca] gradient. All these results indicate that tomato pollen tube is an excellent model to study calcium dynamics.

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Roles of three Fusarium graminearum membrane Ca2+ channels in the formation of Ca2+ signatures, growth, development, pathogenicity and mycotoxin production

Kim

Son

et al. 2018

Fungal Genetics and Biology

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Cytosolic free calcium dynamics as related to hyphal and colony growth in the filamentous fungal pathogen Colletotrichum graminicola

Cited by 8 publications

References 55 publications

The Transient Receptor Potential (TRP) Channel Family in Colletotrichum graminicola: A Molecular and Physiological Analysis

The Transient Receptor Potential (TRP) Channel Family in Colletotrichum graminicola: A Molecular and Physiological Analysis

Calcium dynamics in tomato pollen tubes using the Yellow Cameleon 3.6 sensor

Roles of three Fusarium graminearum membrane Ca2+ channels in the formation of Ca2+ signatures, growth, development, pathogenicity and mycotoxin production

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