Effects of the F-actin inhibitor latrunculin A on the budding yeast Saccharomyces cerevisiae

Kopecká, Marie; Yamaguchi, Masashi; Kawamoto, Susumu

doi:10.1099/mic.0.000091

Cited by 9 publications

(3 citation statements)

References 50 publications

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“…Actin cables serve as tracks to transport mRNAs, proteins, mitochondria, and ribosomes from the mother cell to the daughter cell. Actin patches are clustered in the secretion and endocytosis sites for critical roles in plasma membrane invagination during endocytosis [37] and cell wall remodeling through orienting the polarized secretion of cell wall constituents and enzymes towards the actin patches, promoting bud emergence [38, 39].…”

Section: Resultsmentioning

confidence: 99%

Chemogenomic profiling to understand the antifungal action of a bioactive aurone compound

et al. 2019

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Every year, more than 250,000 invasive candidiasis infections are reported with 50,000 deaths worldwide. The limited number of antifungal agents necessitates the need for alternative antifungals with potential novel targets. The 2-benzylidenebenzofuran-3-(2H)-ones have become an attractive scaffold for antifungal drug design. This study aimed to determine the antifungal activity of a synthetic aurone compound and characterize its mode of action. Using the broth microdilution method, aurone SH1009 exhibited inhibition against C. albicans, including resistant isolates, as well as C. glabrata, and C. tropicalis with IC50 values of 4–29 μM. Cytotoxicity assays using human THP-1, HepG2, and A549 human cell lines showed selective toxicity toward fungal cells. The mode of action for SH1009 was characterized using chemical-genetic interaction via haploinsufficiency (HIP) and homozygous (HOP) profiling of a uniquely barcoded Saccharomyces cerevisiae mutant collection. Approximately 5300 mutants were competitively treated with SH1009 followed by DNA extraction, amplification of unique barcodes, and quantification of each mutant using multiplexed next-generation sequencing. Barcode post-sequencing analysis revealed 238 sensitive and resistant mutants that significantly (FDR P values ≤ 0.05) responded to aurone SH1009. The enrichment analysis of KEGG pathways and gene ontology demonstrated the cell cycle pathway as the most significantly enriched pathway along with DNA replication, cell division, actin cytoskeleton organization, and endocytosis. Phenotypic studies of these significantly enriched responses were validated in C. albicans. Flow cytometric analysis of SH1009-treated C. albicans revealed a significant accumulation of cells in G1 phase, indicating cell cycle arrest. Fluorescence microscopy detected abnormally interrupted actin dynamics, resulting in enlarged, unbudded cells. RT-qPCR confirmed the effects of SH1009 in differentially expressed cell cycle, actin polymerization, and signal transduction genes. These findings indicate the target of SH1009 as a cell cycle-dependent organization of the actin cytoskeleton, suggesting a novel mode of action of the aurone compound as an antifungal inhibitor.

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Section: Resultsmentioning

confidence: 99%

Chemogenomic profiling to understand the antifungal action of a bioactive aurone compound

et al. 2019

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“…Although the relationship between cell size and actin cytoskeleton in yeast is still unclear (Jorgensen et al 2002;Goranov and Amon 2010;Turner et al 2012;Sattlegger et al 2014;Kopecka et al 2015), the connection between the CWI pathway and the actin cytoskeleton (see Introduction) and the rapid shrinking of daughters at the time of cytokinesis suggests that satellite daughters result from an actin cytoskeleton defect. Thus, we asked whether satellite daughters displayed defects in the actin cytoskeleton.…”

Section: Satellite Daughters Display Diffuse Actin Stainingmentioning

confidence: 99%

Shrinking Daughters: Rlm1-Dependent G1/S Checkpoint MaintainsSaccharomyces cerevisiaeDaughter Cell Size and Viability

et al. 2017

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The Rlm1 transcription factor is a target of the cell wall integrity pathway. We report that an Δ mutant grown on a nonfermentable carbon source at low osmolarity forms cell groups in which a mother cell is surrounded by smaller "satellite-daughter" cells. Mother cells in these groups progressed through repeated rounds of cell division with normal rates of bud growth and genetic stability; however, these cells underwent precocious START relative to wild-type mothers. Thus, once activated, Rlm1 delays the transition from G to S, a mechanism we term the cell wall/START (CW/START) checkpoint. The Δ satellite-cell phenotype is suppressed by deletion of either, which encodes the kinase that activates Rlm1, or , which is also activated by Slt2; suggesting that Slt2 can have opposing roles in regulating the START transition. Consistent with an Rlm1-dependent CW/START checkpoint,Δ satellite daughters were unable to grow or divide further even after transfer to rich medium, but UV irradiation in G could partially rescue Δ satellite daughters in the next division. Indeed, after cytokinesis, these satellite daughters shrank rapidly, displayed amorphous actin staining, and became more permeable. As a working hypothesis, we propose that duplication of an "actin-organizing center" in late G may be required both to progress through START and to reestablish the actin cytoskeleton in daughter cells.

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“…Previously, C. neoformans was studied by electron microscopy [3], and the effects of some of these inhibitors were monitored [9,10]. Electron microscopy has also been used for examining inhibition in other yeasts [11,12,13,14,15]. …”

Section: Introductionmentioning

confidence: 99%

Microtubules and Actin Cytoskeleton of Cryptococcus neoformans as Targets for Anticancer Agents to Potentiate a Novel Approach for New Antifungals

Kopecká

2015

Chemotherapy

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Background: We investigated the targeting of microtubules (MT) and F-actin cytoskeleton (AC) of the human pathogenic yeast Cryptococcus neoformans with agents for cancer therapy, in order to examine whether this yeast cytoskeleton could become a new antifungal target for the inhibition of cell division. Methods: Cells treated with 10 cytoskeleton inhibitors in yeast extract peptone dextrose medium were investigated by phase-contrast and fluorescence microscopy, and growth inhibition was estimated by cell counts using a Bürker chamber and measuring absorbance for 6 days. Results: Docetaxel, paclitaxel, vinblastine sulfate salt, cytochalasin D and chlorpropham [isopropyl N-(3-chlorophenyl) carbamate] did not inhibit proliferation. The MT inhibitors methyl benzimidazole-2-ylcarbamate (BCM), nocodazole, thiabendazole (TBZ) and vincristine (VINC) disrupted MT and inhibited mitoses, but anucleated buds emerged on cells that increased in size, vacuolated and seemed to die after 2 days. The response of the cells to the presence of the actin inhibitor latrunculin A (LA) included the disappearance of actin patches, actin cables and actin rings; this arrested budding and cell division. However, in 3-4 days, resistant budding cells appeared in all 5 inhibitors. Disruption of the MT and AC and inhibition of cell division and budding persisted only when the MT and AC inhibitors were combined, i.e. VINC + LA, BCM + LA or TBZ + LA. Conclusion: The MT and AC of C. neoformans are new antifungal targets for the persistent inhibition of cell division by combined F-actin and MT inhibitors.

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Effects of the F-actin inhibitor latrunculin A on the budding yeast Saccharomyces cerevisiae

Cited by 9 publications

References 50 publications

Chemogenomic profiling to understand the antifungal action of a bioactive aurone compound

Chemogenomic profiling to understand the antifungal action of a bioactive aurone compound

Shrinking Daughters: Rlm1-Dependent G1/S Checkpoint MaintainsSaccharomyces cerevisiaeDaughter Cell Size and Viability

Microtubules and Actin Cytoskeleton of Cryptococcus neoformans as Targets for Anticancer Agents to Potentiate a Novel Approach for New Antifungals

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