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|Ph.D., University of Washington and Fred Hutchinson Cancer Research Center, 2004|
Figure 1. The structure of a homodimer of the human septin SEPT2 bound to the nonhydrolyzable GTP analog GppNHp (PDB 3FTQ) with the residues corresponding to those we found in temperature-sensitive yeast rendered as spheres. GppNHp is shown in orange. From Weems, et al GENETICS 2013.
Figure 2. Model for chaperone-mediated quality control of higher-order septin assembly. Nascent septin polypeptides emerging from the ribosome encounter a number of cytosolic chaperones during subsequent de novo folding. Wild-type septins efficiently adopt quasi-native conformations, thereby burying hydrophobic residues and escaping chaperone-mediated sequestration. Heterodimerization with other septins—the first oligomerization step toward septin filament assembly—occurs concomitant with exit from the chamber of the cytosolic chaperonin CCT (also called TRiC). Mutant septins that inefficiently fold the G heterodimerization interface are slower to achieve a conformation allowing chaperone release. Interactions with the prefoldin complex (PFD), the Hsp40 chaperone Ydj1, and the disaggregase Hsp104 are particularly prolonged, leading to a delay in availability of the mutant septin for hetero-oligomerization. From Johnson, et al Mol Biol Cell 2015.
Figure 3. Model for the step-wise assembly of yeast septin hetero-octamers. From Weems & McMurray, eLife 2017.
Figure 4. Model for the defects in prospore membrane (PSM) biogenesis in septin-mutant yeast cells. SPB, spindle pole body; LEP, leading edge protein. From Heasley & McMurray, Molecular Biology of the Cell 2016.
Our research focuses on identifying molecular mechanisms underlying the assembly of macromolecular complexes, with a focus on multisubunit complexes formed by septin proteins. All cellular processes require the function of multisubunit complexes, and while much attention has been given to solving the final structures of such assemblies, comparatively little is known about how individual subunits adopt oligomerization-competent conformations and find their partner subunits in the crowded, dynamic cellular milieu. Below is a summary of the past research from our group.
We first used unbiased genetic screening to find evidence that guanine nucleotide binding by septin proteins plays an entirely structural role during filament assembly, which can be bypassed by specific mutations in a key septin-septin oligomerization interface. Considering that one of these mutations has been found to cause male infertility in humans, these findings have compelling clinical relevance.
We then investigated a phenomenon in which, when cells have a choice between a wild-type and a mutant version of a given septin, mutant septins unable to bind nucleotides are discriminated against during higher-order assembly, despite their ability to function normally under the conditions tested. We discovered that prolonged interaction between nascent mutant polypeptides and cytosolic chaperones results in a kinetic delay in the acquisition by the mutant septin of a conformation competent for oligomerization with other septins. Our findings challenged the dominant paradigm that interactions between chaperones and their clients always promote mutant protein function, and provided a new way of thinking about how chaperone interactions might influence the ability of mutant alleles of proteins to perturb the functions of the wild type. This work was published in Molecular Biology of the Cell in 2015 and in Cell Cycle in 2016.
We went on to develop an entirely new technique for monitoring, for the first time in living cells, the step-wise pathway of assembly of protein complexes, and used it to show how nucleotide-induced conformational changes drive the progression of such an ordered pathway for yeast septins. Moreover, we generated and validated a new model for the role of nucleotide hydrolysis by septins during oligomerization, and found that slow GTPase activity by a key septin subunit drives the formation of distinct hetero-oligomers comprised of alternative subunits. Our “GTPase timer” model predicted that changes in the ratio of GTP to GDP in the cytosol would influence the subunit composition of septin complexes. Our experiments supported this model and further provided a functional context for why septin complex composition is tuned to the metabolic state of the cell.
In budding S. cerevisiae cells, septin complexes are restricted to hetero-octamers, yet in other species hetero-hexamers are also found, in which the "central" subunit corresponding to yeast Cdc10 is bypassed during assembly. It was unknown how
this alternative assembly pathway worked. Building from a serendipitous discovery that the small molecule guanidine hydrochloride (GdnHCl) restores high-temperature growth to cdc10 mutants, we found that hexamer assembly (and Cdc10 bypass) in other
species involves GTPase activity by the Cdc3 homolog. The guanidine group of GdnHCl appears to replace the guanidine group of a key arginine residue that is present in Cdc3 homologs in other species that make Cdc10-less hexamers (including humans),
and is missing from S. cerevisiae Cdc3 and the and Kluyveromyces lactis. Thus the hexamer-vs-octamer decision presumably relies on slow GTPase activity by the septin subunit in the Cdc3 position, just like the choice of terminal subinits
in yeast octamers relies on slow GTPase activity by the septin subunit in the Cdc12 position.
Finally, our work suggests for the first time that GdnHCl can be used to functionally replace "missing" arginine side chains in living cells. Since GdnHCl is an FDA-approved drug for human use, and since arginine is the most commonly mutated amino acid in missense mutations causing human disease, these findings may pave the way for the use of GdnHCl to treat a variety of human genetic diseases. This work, which was done in collaboration with Aurélie Bertin in Paris and Amy Gladfelter's lab at UNC Chapel Hill, was published in eLife in 2020.
We studied the functions of the gametogenesis-specific septin subunits in yeast and determined that they play important and unique roles in guiding the biogenesis of new membrane and cell wall. As septins also play important, but poorly understood, roles in human gametogenesis, these findings are also relevant to human septin function. This work, partly in collaboration with the labs of Jeremy Thorner and Eva Nogales at UC Berkeley, was published in Molecular Biology of the Cell and the Journal of Cell Biology in 2016.
We also collaborated with the lab of Ravi Manjithaya (Jawaharlal Nehru Centre for Advanced Scientific Research, India) to show that yeast septins play an early role in macro-autophagy in starved cells. Given the similarities between the membrane dynamics involved in autophagy and those involved in prospore membrane biogenesis, these findings extend our understanding of septin function in membrane dynamics. This work was published in the Journal of Cell Science in 2018.
Finally, we studied the molecular basis of action of a small molecule (forchlorfenuron, FCF) thought to act specifically on septin filament assembly in non-plant eukaryotes and therefore used by many researchers as a “septin drug”. We found clear evidence of off-target effects, using four different eukaryotic cell types. This work was published in Eukaryotic Cell in 2014.
Our studies challenge the idea that thermodynamics (i.e., the affinities between proteins randomly colliding in solution) is the main driving force of protein complex assembly, and favor largely unappreciated roles for the kinetics of protein folding and cooperativity between protein-protein interactions. We are now focusing our attention on the mechanisms by which septin proteins fold in living cells, and how the insights we gained from the study of mitotic yeast septins apply to septins in other developmental contexts, and to oligomers composed of non-septin proteins.
View Dr. McMurray's Latest Publications in PubMed
Chaperone requirements for de novo folding of Saccharomyces cerevisiae septins. Daniel Hassell, Ashley Denney, Emily Singer, Aleyna Benson, Andrew Roth, Julia Ceglowski, Marc Steingesser, and Michael McMurray. 2022
Proteasome activity modulates amyloid toxicity. Galvin, John; Curran, Elizabeth; Arteaga, Francisco; Goossens, Alicia; Aubuchon-Endsley, Nicki; McMurray, Michael; Moore, Jeffrey; Hansen, Kirk; Chial, Heidi; Potter, Huntington; Brodsky, Jeffrey; Coughlan, Christina. 2022
Meeting report - the ever-fascinating world of septins. Anita Ballet, Michael A. McMurray, Patrick W.
Post-Transcriptional Control of Mating-Type Gene Expression during Gametogenesis in Saccharomyces cerevisiae. Randi Yeager, G. Guy Bushkin, Emily Singer, Rui Fu, Benjamin Cooperman, and Michael McMurray. 2021
Chemical rescue of mutant proteins in living Saccharomyces cerevisiae cells by naturally occurring small molecules. Daniel S Hassell, Marc G Steingesser, Ashley S Denney, Courtney R Johnson, Michael A McMurray. 2021
Discussed in: Naturally occurring small molecules correct mutant proteins in living cells
Selective functional inhibition of a tumor-derived p53 mutant by cytosolic chaperones identified using split-YFP in budding yeast. Ashley S Denney, Andrew D Weems, Michael A McMurray. 2021
Saccharomyces spores are born prepolarized to outgrow away from spore–spore connections and penetrate the ascus wall. Lydia R. Heasley, Emily Singer, Benjamin J. Cooperman, Michael A. McMurray. 2020
Masters of asymmetry - lessons and perspectives from 50 years of septins. Spiliotis ET and McMurray MA. 2020
Guanidine hydrochloride reactivates an ancient septin hetero-oligomer assembly pathway in budding yeast. Johnson CR, Steingesser MG, Weems AD, Khan A, Gladfelter A, Bertin A, McMurray MA. 2020
Turning it inside out: The organization of human septin heterooligomers. McMurray MA and Thorner JT. 2019
The long and short of membrane curvature sensing by septins. McMurray MA. 2019
Septins are involved at the early stages of macroautophagy in S. cerevisiae. Barve G, Sridhar S, Aher A, Sahani MH, Chinchwadkar S, Singh S, K N L, McMurray MA, Manjithaya R. 2018
The step-wise pathway of septin hetero-octamer assembly in budding yeast. Weems A, McMurray M. 2017
Coupling de novo protein folding with subunit exchange into pre-formed oligomeric protein complexes: the 'heritable template' hypothesis. McMurray MA. 2016
Small molecule perturbations of septins. Heasley LR, McMurray MA. 2016
Assays for genetic dissection of septin filament assembly in yeast, from de novo folding through polymerization. McMurray MA. 2016
Kinetic partitioning during de novo septin filament assembly creates a critical G1 "window of opportunity" for mutant septin function. Schaefer RM, Heasley LR, Odde DJ, McMurray MA. 2016
Assembly, molecular organization, and membrane-binding properties of development-specific septins. Garcia G 3rd, Finnigan GC, Heasley LR, Sterling SM, Aggarwal A, Pearson CG, Nogales E, McMurray MA, Thorner J. 2016
Roles of septins in prospore membrane morphogenesis and spore wall assembly in Saccharomyces cerevisiae. Heasley LR, McMurray MA. 2016
Cytosolic chaperones mediate quality control of higher-order septin assembly in budding yeast. Johnson CR, Weems AD, Brewer JM, Thorner J, McMurray MA. 2015
Off-target effects of the septin drug forchlorfenuron on nonplant eukaryotes. Heasley LR, Garcia G 3rd, McMurray MA. 2014
Lean forward: Genetic analysis of temperature-sensitive mutants unfolds the secrets of oligomeric protein complex assembly. McMurray MA. 2014
Higher-order septin assembly is driven by GTP-promoted conformational changes: evidence from unbiased mutational analysis in Saccharomyces cerevisiae. Weems AD, Johnson CR, Argueso JL, McMurray MA. 2014
Native cysteine residues are dispensable for the structure and function of all five yeast mitotic septins. de Val N, McMurray MA, Lam LH, Hsiung CC, Bertin A, Nogales E, Thorner J. 2013
Three-dimensional ultrastructure of the septin filament network in Saccharomyces cerevisiae. Bertin A, McMurray MA, Thorner J, Peters P, Zehr E, McDonald KL, Thai L, Pierson J, Nogales E. 2012
Subunit-dependent modulation of septin assembly: Budding yeast septin Shs1 promotes ring and gauze formation. Garcia G III, Bertin A, Li Z, Song Y, McMurray MA, Thorner J, Nogales E. 2011
Genetic interactions with mutations affecting septin assembly reveal ESCRT functions in budding yeast cytokinesis. McMurray MA, Stefan CJ, Wemmer M, Odorizzi G, Emr SD, Thorner J. 2011
Septin filament formation is essential in budding yeast. McMurray MA, Bertin A, Garcia III G, Lam L, Nogales EE and Thorner J. 2011
Phosphatidylinositol-4,5-bisphosphate promotes budding yeast septin filament assembly and organization. Bertin A, McMurray MA, Thai L, Garcia III G, Votin V, Grob P, Allyn T, Thorner J and Nogales EE. 2010
Pheromone-induced anisotropy in yeast plasma membrane phosphatidylinositol-4,5-bisphosphate distribution is required for MAPK signaling. Garrenton LS, Stefan C, McMurray MA, Emr SD and Thorner J. 2010
Septins: molecular partitioning and the generation of cellular asymmetry. McMurray MA and Thorner J. 2009
Reuse, replace, recycle: Specificity in subunit inheritance and assembly of higher-order septin structures during mitotic and meiotic division in budding yeast. McMurray MA and Thorner J. 2009
Biochemical properties and supramolecular architecture of septin hetero-oligomers and septin filaments. McMurray MA and Thorner J. 2008
Septin stability and recycling during dynamic structural transitions in cell division and development. McMurray MA Thorner J. 2008
Saccharomyces cerevisiae septins: supramolecular organization of heterooligomers and the mechanism of filament assembly. Bertin A*, McMurray MA*, Grob P*, Park SS, Garcia G 3rd, Patanwala I, Ng HL, Alber T, Thorner J, Nogales E. 2008
PhD student (Molecular Biology Program)
Lab Manager (and lab dad)
Professional Research Assistant
PhD student (Molecular Biology Program)
PhD student (Structural Biology and Biochemistry Program)
PhD student in the Molecular, Cell, and Development program, University of Colorado Boulder
Ashley Denney, PhD
MD student (MSTP ), University of Colorado AMC
Andrew Roth, PhD
Research Writer and Regulatory Specialist in the Office of the Vice Chancellor for Research, University of Colorado AMC
AAV Research Associate, Rocket Pharmaceuticals
PhD student in the Molecular Biology program, Princeton University
Lydia Heasley, PhD
Assistant Professor in the Department of Biochemistry and Molecular Genetics, University of Colorado AMC
Andrew Weems, PhD
Instructor with the Lyda Hill Department of Bioinformatics and Post-Doc in the Danuser Lab, UT Southwestern Medical Center
Dental School student,University of Colorado AMC
PRA in Gastroenterology, University of Colorado AMC
Christina Coughlan, PhD, FCP, SI, EMT
Research instructor in Department of Neurology, University of Colorado AMC
Funded by the National Science Foundation through award 1928900
After >100 years of study, the budding yeast Saccharomyces cerevisiae is the best understood eukaryotic cell. A key feature that makes S. cerevisiae such a powerful tool for genetical manipulation is the efficient alternation of haploid and diploid phases (Fig. 1). Upon nutrient deprivation, most diploid S. cerevisiae strains undergo meiosis and sporulation, typically producing four haploid spores within each sporulating cell. Each spore is encased in a specialized wall that confers resistance to a variety of environmental stressors. As the two mating types reflect alternative alleles at a single locus, each meiosis produces two pairs of spores of opposite mating types, "a" and "alpha". In the lab we prevent mating between spores by physically separating them before exposing them to nutrients, which allows them to grow out from the spore wall (“germinate”) and proliferate indefinitely via budding (Fig.1). Whereas a haploid spore from most natural isolates is able to switch mating types and, via subsequent mating with one of its offspring, return to the diploid state (Fig. 1), labs use haploid strains incapable of switching. Instead, diploids are made at will by mating between haploids placed in close proximity. The ability to manipulate genes and study effects in the haploid phase and then to combine different alleles via mating/recombination/sporulation is the foundation of yeast genetics. Exploiting this life cycle in the lab context has extended our understanding of the cellular and molecular biology of S. cerevisiae to a level of detail unparalleled by any other eukaryote.
Despite (or, more likely, because of) our focus on S. cerevisiae as a model for human biology, the yeast field tends to ignore the aspects of yeast biology that lack direct counterparts in human cells. Only recently have we begun to realize that in order to fully understand yeast biology, we must consider how this organism lives outside the lab.
Since in the lab germination is almost always done with isolated spores, most yeast researchers assume that if the spores are kept in contact, they will always mate. Not true! Spores frequently bud even when they are right next to a potential mating partner. Why? How does a spore decide whether to mate, or to bud?
We want to understand the circumstances in which a germinating spore buds vs mates. The research goal of this outreach program is to test the hypothesis that the tendency of a natural Saccharomyces isolate to bud vs mate upon germination can be predicted by assessing HO gene function. The HO gene is essential for mating-type switching and is only expressed in haploid cells that have budded at least once (Fig. 1). HO is thought to have evolved to allow a return to diploidy in cases where spores are dispersed. This trait is called homothallism; the failure to do so is heterothallism. If a strain never sporulates or always mates upon germination (no “lonely spores”), HO is never expressed, and mutations can, in principle, accumulate.
We hypothesize that natural Saccharomyces isolates carrying mutant HO alleles but capable of sporulation will be biased towards intra-ascus mating upon germination. To test this hypothesis, we isolate Saccharomyces species from the wild, sequence the HO locus in each, and assess sporulation ability and bud-vs-mate decision upon germination.
To engage the public in scientific research and enrich public school education in a way that also advances the research goals of our project, we involve members of the general public in the isolation and identification of yeast strains from the bark of oak trees or sourdough starters, and the identification of mutations in HO causing defects in mating-type switching. Here we provide a collection of resources that we have developed, which we hope will be of value to other groups undertaking similar outreach efforts.
In October 2019 for the first time we incorporated undergraduate students from Colorado Christian University in the outreach activity with Morey Middle School 6th-grade students. CCU students (Biology and Elementary Education majors) were exposed directly to what it is like to teach Biology to middle-school students. CCU students also received the oak bark samples and successfully isolated yeast from them, including Saccharomyces cerevisiae, which was confirmed by PCR of ITS2 DNA and sequencing. At the time of the COVID-19 pandemic shutdown, CCU students were attempting to amplify the HO gene for sequencing. CCU students thus also received direct, hand-on training in biological research.
|Intro lecture|| Wild Yeast Project.pdf|
|DNA barcoding lecture||Species ID by DNA sequencing.pdf|
|Yeast ID by microscopy quiz|| Wild Yeast Microscopy of Bark Cultures|
|Map of oak yeast identified in Cheesman Park||FinalCheesmanYeastMap.pdf|
The COIVD-19 pandemic closed Morey Middle School and the Aurora School of Science and Technology, preventing the planned Spring 2020 outreach activities. PI Michael McMurray regularly attends an entirely volunteer-led annual retreat for families with profoundly gifted children (PG Retreat, or PGR; www.pgretreat.org). The in-person event scheduled for the end of June 2020 was canceled, and Michael volunteered to help design and execute a set of virtual, online activities in its place. Michael realized that with the explosion of the use and creation of sourdough starters that accompanied the pandemic-associated shelter-in-place orders, a large number of PGR registrants would likely be interested in a version of the Wild Yeast Isolation outreach activity modified to use sourdough starters as a source of yeast.
Michael spoke with scientists from North Carolina State University’s “Public Science Lab” (http://robdunnlab.com), which has been organizing a “Science of Sourdough” citizen science project (http://robdunnlab.com/projects/wildsourdough) that asks citizens from all over the world to measure properties of sourdough starters. Michael then recruited >20 PGR registrants to participate in a 7-week program that involved many of the same activities as used for middle-school students, but also included isolation of the yeast from the starters using homemade medium, and mailing the yeast to the McMurray lab for analysis. We were successful!
|Intro lecture||Wild Sourdough Yeast|
|DNA barcoding lecture||Yeast 23 & Me|
|Home microbiology basics lecture||Isolating Microbes from Starters|