Tools for live-cell imaging of cytoskeletal and nuclear behavior in the unconventional yeast, <i>Aureobasidium pullulans</i>

Petrucco, Claudia A.; Crocker, Alex W.; D’Alessandro, Alec; Medina, Edgar M.; Gorman, Olivia; McNeill, Jessica; Gladfelter, Amy S.; Lew, Daniel J.

doi:10.1091/mbc.e23-10-0388

Cited by 2 publications

References 57 publications

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Chemical transformation of the multibudding yeast, Aureobasidium pullulans

Wirshing,

Petrucco,

Lew

2024

Journal of Cell Biology

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Aureobasidium pullulans is a ubiquitous polymorphic black yeast with industrial and agricultural applications. It has recently gained attention amongst cell biologists for its unconventional mode of proliferation in which multinucleate yeast cells make multiple buds within a single cell cycle. Here, we combine a chemical transformation method with genome-targeted homologous recombination to yield ∼60 transformants/μg of DNA in just 3 days. This protocol is simple, inexpensive, and requires no specialized equipment. We also describe vectors with codon-optimized green and red fluorescent proteins for A. pullulans and use these tools to explore novel cell biology. Quantitative imaging of a strain expressing cytosolic and nuclear markers showed that although the nuclear number varies considerably among cells of similar volume, total nuclear volume scales with cell volume over an impressive 70-fold size range. The protocols and tools described here expand the toolkit for A. pullulans biologists and will help researchers address the many other puzzles posed by this polyextremotolerant and morphologically plastic organism.

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Chemical transformation of the multibudding yeast, Aureobasidium pullulans

Wirshing,

Petrucco,

Lew

2024

Journal of Cell Biology

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Negative feedback equalizes polarity sites in a multi-budding yeast

Crocker,

Petrucco,

Guan

et al. 2024

Preprint

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Morphogenesis in fungi and animals is directed by polarization of small GTPases Cdc42 and Rac. In the budding yeastSaccharomyces cerevisiaecompetition between polarity patches results in one polarized patch and the growth of a single bud. Here, we describe cell polarity in the yeastAureobasidium pullulans, which establishes multiple coexisting polarity sites yielding multiple buds during a single cell division cycle. Polarity machinery components oscillate in their abundance in these coexisting sites but do so independently of one another, pointing to a lack of global coupling between sites. Previous theoretical work has demonstrated that negative feedback in a polarity circuit could promote coexistence of multiple polarity sites, and time-delayed negative feedback is known to cause oscillations. We show that both these features of negative feedback depend on a protein we identified as Pak1, and that Pak1 requires Rac1 but not Cdc42 for its localization. This work shows how conserved signaling networks can be modulated for distinct morphogenic programs even within the constraints of fungal budding.

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Tools for live-cell imaging of cytoskeletal and nuclear behavior in the unconventional yeast, Aureobasidium pullulans

Cited by 2 publications

References 57 publications

Chemical transformation of the multibudding yeast, Aureobasidium pullulans

Chemical transformation of the multibudding yeast, Aureobasidium pullulans

Negative feedback equalizes polarity sites in a multi-budding yeast

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