Summary

Introduction

We study questions in fundamental cell biology, using fungal models and a mix of experimental and computational approaches. Fungi and animals share conserved molecular strategies to perform many core cell functions, so the tractable yeast Saccharomyces cerevisiae provides a superb model system to gain in-depth understanding that can be translated into computational models. We also study an emerging non-model fungus, Aureobasidium pullulans, that is an ubiquitous poly-extremophile with unconventional growth modes that raise novel questions in cell biology.

Some Questions of Interest

• How do cells regulate cell polarity to achieve different morphologies?

•  How do cells orient cell polarity in response to extracellular signals?

•  How do cells distribute their contents, particularly in complex geometries?

• How do fungi growing under stringent turgor pressure expand their cell walls without lysing?

•  How do cell-cell contacts between cell walls communicate mechanical information to the cell?

Approach

We address these questions using a combination of genetic, cell biological, and computational approaches. Imaging of various cells (including mutants based on prior knowledge) in different conditions with high spatio-temporal resolution often suggests hypotheses that can be tested by generating other informative mutants. Conceptual models can be translated into computational models to test their plausibility, which can also lead to ideas that can be tested experimentally. By developing custom image analysis tools, we can quantify behaviors for more precise matching between model and experiment.

Cell Polarity: Number and stability of polarity sites

Diverse cell types rely on a common molecular pathway, centered on the conserved GTPase Cdc42, to establish cell polarity and develop cell shapes and contacts that are critical for cell function. Polarization signals act through Cdc42-directed regulators to promote accumulation of membrane-bound active GTP-Cdc42 at the site destined to become the cell’s "front". Cdc42 then organizes cytoskeletal elements through a variety of “effectors”, proteins that bind specifically to GTP-Cdc42. Our work identified a positive feedback mechanism that promotes concentration of active Cdc42 to form a polarity site. Positive feedback concentrates active Cdc42 at a single cortical site, which is critical for budding of yeast cells and directed migration of animal cells. But some cells generate several polarity sites to make more complex morphologies. How is the number of sites encoded in the polarity circuit?  Also, when cells need to respond to external cues, they can undergo a search process where they rapidly assemble and disassemble polarity sites in different places. Computational explorations indicate that it is very difficult for a polarity circuit designed to form a stable front to exhibit this searching behavior. So how do cells do it?

Reviews: Wu & Lew 2013, Chiou et al. 2017.

Orienting Polarity: How yeast cells find their mates

Polarization is often oriented by physical or chemical features or the cell’s environment, and cells are extraordinarily good at detecting and decoding chemical gradients in noisy and complex environments. Micro-organisms track gradients to find food or mates, and gradient detection underlies axon guidance, homing of immune cells towards invaders, chemotaxis of fibroblasts towards wound sites, and guidance of sperm towards the egg. To track a chemical gradient, eukaryotic cells compare concentrations at different points on the cell’s surface. Gradient decoding becomes most challenging when cells are small (so concentrations can only be compared across short distances), when gradients are shallow (so that concentrations differ very little across a short distance), and when chemical concentrations are low (so that molecular noise is significant). Also challenging are situations when there is more than one source of attractant. Yeast cells seeking mates encounter a combination of ALL these challenges, yet remarkably they can decode gradients of pheromones to find mates. How do they perform this astonishing feat?

Reviews: Clark-Cotton et al. 2022, Ghose et al. 2022.

Mechanobiology of the fungal cell wall

Fungal infections are on the rise, and climate change is predicted to acclimate fungi to human body temperature, potentially creating a surge in fungal disease that we are ill-equipped to treat, because (unlike bacteria) fungi and animals share most of their cell biology. However, a major difference between animals and fungi, and a potential Achilles’ heel, is the fungal cell wall. Fungal cells live in conditions that drive water influx into the cell, leading to an expansion that would catastrophically lyse the cell if it were not protected by a rigid cell wall. The cell wall is composed of a mesh of cross-linked carbohydrate polymers and glycoproteins. In order to grow, this mesh must expand, which involves local wall thinning and remodeling near the polarity site. Small imbalances in wall expansion would easily burst the cells, except that fungi have developed a wall mechanosensing pathway that detects local flaws and rapidly repairs them. How does this pathway detect flaws? How does it pause growth to allow repair? And how do cells silence this pathway when it is safe to remove the cell wall, as happens at contact sites between mating cells?

Reviews: This is a new direction for the lab. Stay tuned!

Cell biology of an unconventional fungus

Tractable yeast model systems have provided much of our molecular understanding of eukaryotic cell biology, because the tools developed to allow rapid and definitive experiments were for decades only available in those systems. However, recent advances have made the path from non-model system to tractable system much faster, allowing investigation into a growing menagerie of “unconventional” systems. One such system that we are developing is the poly-extremophile Aureobasidium pullulans, which has been found in diverse niches including on many trees and terrestrial plants, in the ocean, in salt lakes, and in the Antarctic. The genome of this fungus reveals a capacity for unusual biochemical reactions, stimulating biotechnological interest. Our interest was stimulated by the finding that the A. pullulans lifestyle differs from that of other known budding yeasts. Unlike diverse well-studied yeasts like the baker’s yeast Saccharomyces cerevisiae, the human pathogens Candida albicans and Cryptococcus neoformans, or the plant pathogen Ustilago maydis, which always produce one daughter cell (“bud”) per cell cycle, A. pullulans can produce several buds (>10) simultaneously. This makes A. pullulans a powerful tool for comparative cell biology. How does A. pullulans decide how many buds to make in a given cell cycle? How does it apportion its many nuclei among the buds? How does it distribute organelles between the buds? Are there accurate partitioning systems? If so, how do they work? If not, how do A. pullulans cells tolerate or adjust to variable organelle numbers?

Reviews: This is a new direction for the lab. Stay tuned!

Publications

Daniel Lew's publications on PubMed can be found here.

2024

Petrucco CA, Crocker AW, D'Alessandro A, Medina EM, Gorman O, McNeill J, Gladfelter AS, Lew DJ. Tools for live-cell imaging of cytoskeletal and nuclear behavior in the unconventional yeast, Aureobasidium pullulans. Mol Biol Cell. 2024 Apr 1;35(4):br10. doi: 10.1091/mbc.E23-10-0388. Epub 2024 Mar 6. PMID: 38446617.

2023

Guan K, Curtis ER, Lew DJ, Elston TC (2023) Particle-based simulations reveal two positive feedback loops allow relocation and stabilization of the polarity site during yeast mating. PLOS Computational Biology 19(10): e1011523. https://doi.org/10.1371/journal.pcbi.1011523

2022

Jacobs KC , Gladfelter AS, Lew DJ. Targeted secretion: Myosin V delivers vesicles through formin condensates. Curr Biol. Nov 7;32(21):R1228-R1231 (2022)

Jacobs KC, Gorman O, Lew DJ. Mechanism of commitment to a mating partner in Saccharomyces cerevisiae. Mol Biol Cell. Oct 1;33(12):ar112. (2022)

Jones DS, Gillette DD, Cooper PE, Salinas RY, Hill JL, Black SJ, Lew DJ, Canelas DA. Cultivating PhD aspirations during college. CBE Life Sci Educ. 21:ar22. (2022)

Ghose D, Elston T, Lew D. Orientation of cell polarity by chemical gradients. Annu Rev Biophys. 51:431-451. (2022)

Jacobs KC, Lew DJ. Pheromone guidance of polarity site movement in yeast. Biomolecules. 12(4):502. (2022)

Clark-Cotton MR, Jacobs KC, Lew DJ. Chemotropism and cell-cell fusion in fungi. Microbiol Mol Biol Rev. 86(1):e0016521. (2022)

2021

Ramirez SA, Pablo M, Burk S, Lew DJ, Elston TC. A novel stochastic simulation approach enables exploration of mechanisms for regulating polarity site movement. PLoS Comput Biol. 17(7):e1008525. (2021)

Ghose D, Jacobs K, Ramirez S, Elston T, Lew D. Chemotactic movement of a polarity site enables yeast cells to find their mates. Proceedings of the National Academy of Sciences. 118(22). (2021)

Video abstract (Katherine Jacobs):

Clark-Cotton MR, Henderson NT, Pablo M, Ghose D, Elston TC, Lew DJ. Exploratory polarization facilitates mating partner selection in Saccharomyces cerevisiae. Molecular biology of the cell. 32(10):1048-1063. (2021).

Chiou JG, Moran KD, Lew DJ. How cells determine the number of polarity sites. Elife. 10:e58768. (2021)

Robertson CG, Clark-Cotton MR, Lew DJ. Mechanisms that ensure monogamous mating in Saccharomyces cerevisiae. Molecular Biology of the Cell. 32(8):638-44.  (2021). 

2020

Ghose D and Lew D. Mechanistic insights into actin-driven polarity site movement in yeast. Molecular biology of the cell 31(10):1085-1102. (2020).

Moran KD, Lew DJ. How Diffusion Impacts Cortical Protein Distribution in Yeasts. Cells 9(5):1113. (2020). 

Ramirez, SA., Michael P, Sean B, Lew DJ, and Elston T. A Novel Stochastic Simulation Approach Enables Exploration of Mechanisms to Regulate Polarization Dynamics. Biophysical Journal 118, no. 3: 135a. (2020).

2019

Henderson NT, Pablo M, Ghose D, Clark-Cotton MR, Zyla TR, Nolen J, Elston TC, Lew DJ. Ratiometric GPCR signaling enables directional sensing in yeast. PLoS Biol. 17(10):e3000484. (2019).

Mitchison-Field LMY, Vargas-Muñiz JM, Stormo BM, Vogt EJD, Van Dierdonck S, Pelletier JF, Ehrlich C, Lew DJ, Field CM, Gladfelter AS. Unconventional Cell Division Cycles from Marine-Derived Yeasts. Curr Biol. 29(20):3439-56. (2019).

2018

Daniels, CN, Zyla, TR, and Lew, DJ A role for Gic1 and Gic2 in Cdc42 polarization at elevated temperature. PLoS ONE. (2018). 

Moran KD, Kang H, Araujo AV, Zyla TR, Saito K, Tsygankov D, Lew DJ. Cell-cycle control of cell polarity in yeast. J Cell Biol 218: 171-189. (2018).

McClure, A.W., Jacobs, K.C., Zyla, T.R., and Lew, D.J. Mating in wild yeast: delayed interest in sex after spore germination. Mol. Biol. Cell. (2018). 

Lai H, Chiou JG, Zhurikhina A, Zyla TR, Tsygankov D, Lew DJ. Temporal regulation of morphogenetic events in Saccharomyces cerevisiae. Mol Biol Cell 29: 2069-2083. (2018).

Chiou JG, Ramirez SA, Elston TC, Witelski TP, Schaeffer DG, Lew DJ. Principles that govern competition or co-existence in Rho-GTPase driven polarization. PLoS Comput. Biol. 14(4): e1006095 (2018).

2017

Chiou JG, Balasubramanian MK, Lew DJ. Cell Polarity in Yeast. Annu. Rev. Cell. Dev. Biol. 33:77-101. (2017).

Woods B, Lew DJ. Polarity establishment by Cdc42: Key roles for positive feedback and differential mobility. Small GTPases. 28:1-8. (2017).

2016

Woods B, Lai H, Wu CF, Zyla TR, Savage NS, Lew DJ. Parallel Actin-Independent Recycling Pathways Polarize Cdc42 in Budding Yeast. Curr Biol. 22;26(16):2114-26. doi: 10.1016/j.cub.2016.06.047. (2016).

Kang H, Lew DJ. How do cells know what shape they are? Curr Genet. 63(1):75-77. (2016).

McClure AW, Wu CF, Johnson SA, Lew DJ. Imaging Polarization in Budding Yeast. Methods Mol Biol. 1407:13-23. (2016).

Kang, H., Tsygankov, D., and Lew, D.J. Sensing a bud in the yeast morphogenesis checkpoint: a role for Elm1. Mol. Biol. Cell 27: 1764-1775 (2016).

2015

McClure A.W., Minakova M., Dyer J.M., Zyla T.R., Elston T.C., Lew D.J. Role of Polarized G Protein Signaling in Tracking Pheromone Gradients. Dev. Cell35(4):471-82 (2015).

Wu C.F., Chiou J.G., Minakova M., Woods B., Tsygankov D., Zyla T.R., Savage N.S., Elston T.C., Lew D.J. Role of competition between polarity sites in establishing a unique front. Elife 4:e11611 (2015).

Woods B., Kuo C.C., Wu C.F., Zyla T.R., Lew D.J. Polarity establishment requires localized activation of Cdc42. J. Cell Biol. 211(1):19-26 (2015).

Ramirez S.A., Raghavachari S., Lew D.J. Dendritic spine geometry can localize GTPase signaling in neurons. Mol. Biol. Cell 26(22):4171-81 (2015).

McClure A.W., Lew D.J. To avoid a mating mishap, yeast focus and communicate. J. Cell Biol. 208(7):867-868 (2015).

2014

McClure A.W., Lew D.J. Cell polarity: netrin calms an excitable system. Current Biology 24(21):R1050-2 (2014).

Kuo C.C., Savage N.S., Chen H., Wu C.F., Zyla T.R., Lew D.J. Inhibitory GEF phosphorylation provides negative feedback in the yeast polarity circuit. Current Biology 24(7):753-9 (2014).

2013

Wu C.F., Lew D.K. Beyond symmetry-breaking: competition and negative feedback in GTPase regulation Trends in Cell Biology 23(10):476-83 (2013). *Top ten editorial board favorite article of 2013

Wu C.F., Savage N.S., Lew D.J. Interaction between bud-site selection and polarity-establishment machineries in budding yeast Philosophical Transactions of the Royal Society B 368(1629) (2013).

King K., Kang H., Jin M., Lew D.J. Feedback control of Swe1p degradation in the yeast morphogenesis checkpoint Molecular Biology of the Cell 24(7):914-22 (2013).

Dyer J.M., Savage N.S., Jin M., Zyla T.R., Elston T.C., Lew D.J. Tracking shallow chemical gradients by actin-driven wandering of the polarization site Current Biology 23(1):32-41 (2013).

2012

King K., Jin M., Lew D. Roles of Hsl1p and Hsl7p in Swe1p degradation: beyond septin tethering Eukaryotic Cell 11(12):1496-502 (2012).

Chen H., Kuo C.C., Kang H., Howell A.S., Zyla T.R., Jin M., Lew D.J. Cdc42p regulation of the yeast formin Bni1p mediated by the effector Gic2p Molecular Biology of the Cell 23(19):3814-26 (2012).

Howell A.S., Jin M., Wu C.F., Zyla T.R., Elston T.C., Lew D.J. Negative feedback enhances robustness in the yeast polarity establishment circuit Cell149(2):322-33 (2012).

Savage N.S., Layton A.T., Lew D.J. Mechanistic mathematical model of polarity in yeast Molecular Biology of the Cell 23(10):1998-2013 (2012).

Howell A.S., Lew D.J. Morphogenesis and the cell cycle Genetics 190(1):51-77 (2012).

2011

Johnson J.M., Jin M., Lew D.J. Symmetry breaking and the establishment of cell polarity in budding yeast Current opinion in genetics & development 21(6):740-6 (2011).

Chen H., Howell A.S., Robeson A., Lew D.J. Dynamics of septin ring collar formation in Saccharomyces cerevisiae Biological Chemistry 392(8-9):689-97 (2011).

Layton A.T., Savage N.S., Howell A.S., Carroll S.Y., Drubin D.G., Lew D.J. Modeling vesicle traffic reveals unexpected consequences for Cdc42p-mediated polarity establishment Current Biology 21(3): 184-94 (2011).

2009

Howell A.S., Savage N.S., Johnson S.A., Bose I., Wagner A.W., Zyla T.R., Nijhout H.F., Reed M.C., Goryachev A.B., and Lew D.J. Singularity in Polarization: Rewiring Yeast Cells to Make Two Buds Cell 139(4):731-43 (2009).

Crutchley J., King K.M., Keaton M.A., Szkotnicki L., Orlando D.A., Zyla T.R., Bardes E.S., and Lew D.J. Molecular dissection of the checkpoint kinase Hsl1p.Molecular Biology of the Cell 20(7):1926-36 (2009).

Kozubowski L., Saito K., Johnson J.M., Howell A.S., and Lew D.J. Response: GEF localization, not just activation, is needed for yeast polarity establishment. Current Biology 19(5):R195 (2009).

2008

Kozubowski L., Saito K., Johnson J.M., Howell A.S., Zyla T.R., and Lew D.J. Symmetry-breaking polarization driven by a Cdc42p GEF-PAK complex. Current Biology 18(22):1719-26 (2008).

Szkotnicki L., Crutchley J.M., Zyla T.R., Bardes E.S., and Lew D.J. The checkpoint kinase Hsl1p is activated by Elm1p-dependent phosphorylation. Molecular Biology of the Cell 19(11):4675-86 (2008).

Keaton M.A., Szkotnicki L., Marquitz A.R., Harrison J., Zyla T.R., and Lew D.J. Nucleocytoplasmic trafficking of G2/M regulators in yeast. Molecular Biology of the Cell 19(9):4006-18 (2008).

Lew, D.J., Burke, D.J., and Dutta, A. The immortal strand hypothesis: how could it work? Cell 133: 21-23 (2008).

York, J.D. and Lew, D.J. IP7 guards the CDK gate. Nature Chem. Biol. 4: 16-17 (2008).

2007

Tong, Z., Gao, X-G., Howell, A., Bose, I., Lew, D.J., and Bi, E. Adjacent positioning of cellular structures enabled by a Cdc42 GAP mediated zone of inhibition.J. Cell Biol. 7: 1375-1384 (2007).

Keaton, M., Bardes, E.S.G., Marquitz, A.R., Freel, C.D., Zyla, T.R., Rudolph, J., and Lew, D.J. Differential susceptibility of S and M phase cyclin/CDK complexes to inhibitory tyrosine phosphorylation in yeast. Current Biology 17: 1181-1189 (2007).

Haase, S.B., and Lew, D.J. Microtubule Organization: Cell Fate is Destiny. Current Biology r248-r251 (2007).

2006

Keaton, M., and Lew, D.J. The Morphogenesis Checkpoint: Progress and Controversy. Curr. Opin. Microbiol. 9: 540-546. (2006).

2005

Lew, D.J. Cell Polarity: Negative Feedback Shifts the Focus. Current Biology 15: R994-R996 (2005).

McNulty, J.J., and Lew, D.J. Swe1p responds to cytoskeletal perturbation, not bud size, in S. cerevisiae. Current Biology 15: 2190-2198 (2005).

Gladfelter, A.S., Kozubowski, L., Zyla, T.R., and Lew, D.J. Interplay between septin organization, cell cycle and cell shape in yeast. J. Cell Sci. 118: 1617-1628 (2005).

Irazoqui, J.E., Howell, A.S., Theesfeld, C.L., and Lew,D.J. Opposing roles for actin in Cdc42p polarization. Mol. Biol. Cell 16: 1296-1304 (2005).

2004

Gladfelter, A.S., Zyla, T.R., and Lew, D.J. Genetic interactions among regulators of septin organization. Euk. Cell, 3: 847-854 (2004).

Irazoqui, J.E., Gladfelter, A.S., and Lew, D.J. Cdc42p, GTP hydrolysis, and the cell's sense of direction. Cell Cycle, 3: e53-e56 (2004).

Harrison, J.C., Zyla, T.R., Bardes, E.S.G., and Lew, D.J. Stress-specific activation mechanisms for the "cell integrity" MAPK pathway. J. Biol. Chem., 279: 2616-2622 (2004).

Irazoqui, J.E. and Lew, D.J. Polarity establishment in yeast (Review). J. Cell Sci. 117, 2169-2171 (2004).

2003

Lew, D.J. The Morphogenesis Checkpoint. Curr. Opin. Cell Biol., 15: 648-653. (2003).

Irazoqui, J.E., Gladfelter, A.S., and Lew, D.J. Scaffold-mediated symmetry breaking by Cdc42p. Nature Cell Biology, 5:1062-1070 (2003).

Lew, D.J. and Burke, D.J. The spindle assembly and spindle position checkpoints. Ann. Rev. Genet., 37:251-282 (2003).

Theesfeld, C.L., Zyla, T.R., Bardes, E.S., and D.J. Lew. A monitor for bud emergence in the yeast morphogenesis checkpoint. Mol Biol Cell, 14:3280-3291. (2003).

2002

Gladfelter, A.S., I. Bose, T.R. Zyla, E.S. Bardes, and D.J. Lew Septin ring assembly involves cycles of GTP loading and hydrolysis by Cdc42p. J Cell Biol.156:315-26. (2002).

Lew, D.J.: Formin' actin filament bundles (News & Views). Nature Cell Biol. 4: E29-E30.(2002).

Marquitz, A.R., J.C. Harrison, I. Bose, T.R. Zyla, J.N.McMillan, and D.J. Lew: The Rho-GAP Bem2p plays a GAP-independent role in the morphogenesis checkpoint. EMBO J, 21:4012-4025. (2002).

McMillan, J.N., C.L. Theesfeld, J.C. Harrison, E.S. Bardes, and D.J. Lew. Determinants of Swe1p Degradation in Saccharomyces cerevisiae. Mol Biol Cell, 13:3560-3575. (2002).

2001

Adamo, J.E., Moskow, J.J., Gladfelter, A.S., Viterbo, D., Lew, D.J., and Brennwald, P.J.: Yeast Cdc42 functions at a late step in exocytosis, specifically during polarized growth of the emerging bud. J. Cell Biol. 155: 581-592. (2001).

Gladfelter, A.S., Pringle, J.R., and Lew, D.J.: The septin cortex at the yeast mother-bud neck. Curr. Opin. Microbiol. 4: 681-689. (2001).

Gladfelter, A. S., Moskow, J. J., Zyla, T. R., and Lew, D. J.: Isolation and characterization of effector-loop mutants of CDC42 in yeast. Mol. Biol. Cell, 12: 1239-1255. (2001).

Lew, D.J.: The Cell Cycle. Encyclopedia of Genetics (Sydney Brenner, Ed.), p.286-296. Academic Press. (2001).

Bose, I., Irazoqui, J.E., Moskow, J.J., Bardes, E.S.G., Zyla, T.R., and Lew, D.J.: Assembly of scaffold-mediated complexes containing Cdc42p, the exchange factor Cdc24p, and the effector Cla4p required for cell cycle regulated phosphorylation of Cdc24p. J. Biol. Chem. 276: 7176-7186. (2001).

Harrison, J.C., Bardes, E. S. G., and Lew, D. J.: A role for the Pkc1p/Mpk1p kinase cascade in the morphogenesis checkpoint. Nature Cell Biol. 3: 417-420. (2001).

2000

Yeh, E., Yang, C., Maddox, P., Chin, E., Salmon, E.D., Lew, D.J., and Bloom, K.: Dynamic positioning of mitotic spindles in yeast: role of mitotic motors and asymmetric determinants. Mol. Biol. Cell 11, 3949-3961 (2000).

Moskow, J. J., Gladfelter, A. S., Lamson, R. E., Pryciak, P. M., and Lew, D. J.: The role of Cdc42p in pheromone-stimulated signal transduction in Saccharomyces cerevisiae. Mol. Cell. Biol. 20, 7559-7571. (2000).

Longtine, M. S., Theesfeld, C. L., McMillan, J. N., Weaver, E., Pringle, J. R. and Lew, D. J.: Septin-dependent Assembly of a Cell-cycle-regulatory Module in Saccharomyces cerevisiae. Mol. Cell. Biol., 4049-4061. (2000).

Lew, D.J. Cell-cycle checkpoints that ensure coordination between nuclear and cytoplasmic events in Saccharomyces cerevisiae. Curr. Opin. Genet. Develop. 10, 47-53 (2000).