Fellow

Thomas Lyons

Education

• BA in Chemistry. Middlebury College, Middlebury, VT (1993)
• PhD in Chemistry and Biochemistry. University of California, Los Angeles (1998)
• Postdoctoral Fellow. University of Missouri, Columbia (1998-2001)
• Assistant Professor. University of Florida, Gainesville (2002-2008)
• Principal Research Scientist. Evolugate LLC, Gainesville, FL (2008-present)

Research summary

The Lyons group has pioneered research into the basic chemistry and biology of a novel family of membrane bound receptors known as PAQRs. This superfamily of proteins is nearly ubiquitous in biology being found in all almost Eukaryotes and Eubacteria, but being conspicuously absent in Archaea. The PAQR superfamily can be divided into three classes based on sequence comparisons. Class I receptors are found in nearly all eukaryotes. The two best-characterized Class I receptors are the human adiponectin receptors, AdipoR1 (PAQR1) and AdipoR2 (PAQR2), which sense adiponectin, an adipose-derived hormone in the C1q/TNF superfamily of animal proteins. A third Class I PAQR (Izh2p) from the fungus Saccharomyces cerevisiae senses plant proteins in the PR-5 defensin superfamily whose three-dimensional fold is nearly identical to the β-sandwich of the C1q/TNF superfamily. Consequently, it appears as though the unifying feature of Class I PAQRs may be the ability to bind proteins with a specific β-sandwich fold.

On the other hand, the Class II PAQRs, including human mPRα (PAQR7), mPRβ (PAQR8) and mPRγ (PAQR5) diverged from Class I receptors after the evolution of metazoans. Intriguingly, the steroid hormone progesterone agonizes these receptors and it is unclear how this functionality evolved from Class I β-sandwich receptors. Finally, there is the enigma of the Class III PAQRs, which have the deepest evolutionary roots but no known agonist. Not only do all metazoans have at least one Class III protein, they are widely, but not universally, dispersed in protozoan and eubacterial proteomes. Because Class III receptors predate Class I receptors, one might predict that their most recent common ancestor may have sensed a β-sandwich-like protein.

Receptors in the PAQR family include medically relevant receptors for human hormones. Adiponectin is a recently discovered hormone that plays a critical role in energy metabolism and hypoadiponectinemia may play a central role in the etiology of a variety of obesity-related pathologies, including type II diabetes. Progesterone is best known as a steroid hormone that regulates vertebrate reproduction, however, the true breadth of the physiology of this hormone is only beginning to be discovered.

Important discoveries and interests include:

• Elucidating the mechanism through which PAQR receptors convert extracellular signals into intracellular second messengers. By comparing the sequences of PAQRs to other known proteins, we found that PAQR receptors bear distant similarity to a family of sphingolipid hydrolase enzymes called alkaline ceramidases. Indeed, we discovered that PAQRs produce sphingolipid second messengers that are responsible for physiological changes elicited by activated receptors.
• We discovered that fungal PAQR receptors likely regulate the developmental program called filamentation in response to proteins produced by the plant innate immune system. This is a new paradigm for plant-microbe interactions. It is also critical because these plant proteins are known to survive the fermentation process and could be critical regulators for fungal growth in industrial processes such as the conversion biomass to biofuel.
• We identified the human cytokine tumor necrosis factor alpha (TNFa) as an antagonist of all human PAQR receptors. TNFα also belongs to the C1q/TNF superfamily of vertebrate proteins indicating that all PAQR receptors studied to date actually sense β-sandwich proteins as either agonists or antagonist, thus functionally unifying the entire PAQR family.
• We developed a high throughput yeast-based assay system to screen for modulators of PAQR receptors from any source. We have already begun studying human, frog, ascidian, flatworm, cnidarian, fungal and eubacterial PAQRs. We are interested in the evolution of progesterone as a functional agonist for Class II receptors and in studying the effect of environmental contaminants on progesterone receptors.

As a postdoctoral fellow, Dr. Lyons studied the biochemistry and molecular biology of metal ion homeostasis in fungi. Fungi are important medical and agricultural pests as well as being important for industrial applications. They are also successfully used as a model system for higher eukaryotes. Dr. Lyons studied how these organisms obtain sense nutrient deficiencies, obtain nutrients from the environment and properly metabolize these nutrients. Research focused on zinc homeostasis because zinc is an essential cofactor approximately 10% of eukaryotic proteins in addition to being an important signaling molecule.

Important discoveries include:

• We used transcriptional profiling to screen an entire fungal genome for genes whose transcription responds to changes in the bioavailability of zinc. We found that many zinc-regulated genes were not directly involved in zinc homeostasis - but were rather involved in global metabolic adaptation to zinc-deficiency.
• We began the functional characterization of a variety of zinc-regulated genes of unknown function. This included fungal members of the PAQR family.

As a graduate student, Dr. Lyons studied the biochemistry of metalloproteins with a focus on how metal ions affect the function and stability of copper, zinc superoxide dismutase (SOD), an essential antioxidant enzyme. Dr. Lyons also studied how mutations in this enzyme affect activity and stability. This was a critical endeavor because mutations in this enzyme cause the fatal neurodegenerative disorder, amyotrophic lateral sclerosis by a yet-to-be-identified toxic gain of function. This enzyme is also essential for aerobic growth in microorganisms and is important for maximal life span.

Important discoveries include:

• Our research indicated that the zinc-binding site of SOD was exquisitely sensitive to perturbations in protein structure and that zinc was an important restrictor of enzyme activity. Perturbations of zinc binding could lead to toxic gain of function in mutations that cause ALS.
• We also discovered that SOD from fungi has an inherent asymmetry in the functional dimer and that this asymmetry may be related to the ability of cells to properly distribute copper and zinc to this enzyme.

Awards and Honors

• National Institutes of Health Chemistry-Biology Interface Training Grant Awardee, Graduate Student/Research Assistant, 1994-1996
• University of Missouri Molecular Biology Program, Postodoctoral Fellowship, 1998-2000
• National Institutes of Health/National Research Service Award, Postdoctoral Fellowship, 2000-2001
• Anderson Scholar /University of Florida College of Liberal Arts and Sciences Faculty Honoree. For outstanding undergraduate mentoring, 2004
• Courtesy Appointment, University of Florida, Department of Chemistry, 2009-present

Recent Publications

Non-genomic progesterone signalling and its non-canonical receptor
Moussatche P, Lyons TJ
Biochem Soc Trans (2012) Feb;40(1):200-4
<Abstract>

The steroid hormone progesterone regulates many critical aspects of vertebrate physiology. The nuclear receptor for progesterone functions as a ligand-activated transcription factor, directly regulating gene expression. This type of signalling is referred to as the 'genomic' pathway. Nevertheless, progesterone also stimulates rapid physiological effects that are independent of transcription. This pathway, termed 'non-genomic', is mediated by the mPRs (membrane progesterone receptors). These mPRs belong to a larger class of membrane receptors called PAQRs (progestin and adipoQ receptors), which include receptors for adiponectin in vertebrates and osmotin in fungi. mPRs have been shown to activate inhibitory G-proteins, suggesting that they act as GPCRs (G-protein-coupled receptors). However, PAQRs do not resemble GPCRs with respect to topology or conserved sequence motifs. Instead, they more closely resemble proteins in the alkaline ceramidase family and they may possess enzymatic activity. In the present paper, we highlight the evidence in support of each model and what is currently known for PAQR signal transduction of this non-canonical receptor.

Directed evolution of a filamentous fungus for thermotolerance
de Crecy, E; Jaronski, S; Lyons, B; Lyons, TJ; Keyhani, NO
BMC Biotechnology 9 74 (2009)
<Abstract>

Background: Filamentous fungi are the most widely used eukaryotic biocatalysts in industrial and chemical applications. Consequently, there is tremendous interest in methodology that can use the power of genetics to develop strains with improved performance. For example, Metarhizium anisopliae is a broad host range entomopathogenic fungus currently under intensive investigation as a biologically based alternative to chemical pesticides. However, it use is limited by the relatively low tolerance of this species to abiotic stresses such as heat, with most strains displaying little to no growth between 35-37 degrees C. In this study, we used a newly developed automated continuous culture method called the Evolugator(TM) which takes advantage of a natural selection-adaptation strategy, to select for thermotolerant variants of M. anisopliae strain 2575 displaying robust growth at 37 degrees C. Results: Over a 4 month time course, 22 cycles of growth and dilution were used to select 2 thermotolerant variants of M. anisopliae. Both variants displayed robust growth at 36.5 degrees C, whereas only one was able to grow at 37 degrees C. Insect bioassays using Melanoplus sanguinipes (grasshoppers) were also performed to determine if thermotolerant variants of M. anisopliae retained entomopathogenicity. Assays confirmed that thermotolerant variants were, indeed, entomopathogenic, albeit with complex alterations in virulence parameters such as lethal dose responses (LD50) and median survival times (ST50). Conclusion: We report the experimental evolution of a filamentous fungus via the novel application of a powerful new continuous culture device. This is the first example of using continuous culture to select for complex phenotypes such as thermotolerance. Temperature adapted variants of the insect-pathogenic, filamentous fungus M. anisopliae were isolated and demonstrated to show vigorous growth at a temperature that is inhibitory for the parent strain. Insect virulence assays confirmed that pathogenicity can be retained during the selection process. In principle, this technology can be used to adapt filamentous fungi to virtually any environmental condition including abiotic stress and growth substrate utilization.

Antagonism of Human Adiponectin Receptors and Their Membrane Progesterone Receptor Paralogs by TNF alpha and a Ceramidase Inhibitor
Kupchak, BR; Garitaonandia, I; Villa, NY; Smith, JL; Lyons, TJ
Biochemistry 48 (24) 5504-5506 (2009)
<Abstract>

The progestin and AdipoQ receptor (PAQR) family of proteins comprises three distinct structural classes, each with seemingly different agonist specificities. For example, Class I receptors, like the human adiponectin receptors (AdipoR1 and AdipoR2), sense proteins with a particular three-dimensional fold, while Class II receptors are nonclassical membrane receptors for the steroid hormone progesterone. Using a previously developed heterologous expression system to study PAQR receptor activity, we demonstrate that human PAQRs from all three classes are antagonized by both 1(S),2(R)-D-erythro-2-(N-myristoylamino)-1-phenyl-1-propanol, a ceramidase inhibitor, and TNF alpha, a homologue of adiponectin that functions antagonistically to both adiponectin and progesterone in human cells.

Adiponectin identified as an agonist for PAQR3/RKTG using a yeast-based assay system
Garitaonandia, I; Smith, JL; Kupchak, BR; Lyons, TJ
J. Recept. Signal Transduction 29 (1) 67-73 (2009)
<Abstract>

The PAQR family of proteins comprises an intriguing group of newly discovered receptors. Although the agonist is known for 5 of the 11 human PAQRs, most are considered "orphan" receptors. We developed a yeast-based assay system for PAQR receptor activity that can be used to identify agonists for PAQRs of unknown function. Using this system, we found that the proteinaceous hormone adiponectin functions as an agonist of PAQR3, a previously uncharacterized member of this family. This is not surprising given that PAQR3 is most closely related to PAQR1 (AdipoR1) and PAQR2 (AdipoR2), which also sense adiponectin. The identification of adiponectin as an agonist for PAQR3 is of considerable clinical relevance because adiponectin suppresses the proliferation of tumor cells and it has been reported that PAQR3 suppresses tumorigenesis. Thus, the interaction between PAQR3 and adiponectin may help explain the antiproliferative properties of adiponectin.

Sphingolipids Function as Downstream Effectors of a Fungal PAQR
Villa, NY; Kupchak, BR; Garitaonandia, I; Smith, JL; Alonso, E; Alford, C; Cowart, LA; Hannun, YA; Lyons, TJ
Mol. Pharmacol. 75 (4) 866-875 (2009)
<Abstract>

The Izh2p protein from Saccharomyces cerevisiae belongs to the newly characterized progestin and adipoQ receptor (PAQR) superfamily of receptors whose mechanism of signal transduction is still unknown. Izh2p functions as a receptor for the plant PR-5 defensin osmotin and has pleiotropic effects on cellular biochemistry. One example of this pleiotropy is the Izh2p-dependent repression of FET3, a gene involved in iron-uptake. Although the physiological purpose of FET3 repression by Izh2p is a matter of speculation, it provides a reporter with which to probe the mechanism of signal transduction by this novel class of receptor. Receptors in the PAQR family share sequence similarity with enzymes involved in ceramide metabolism, which led to the hypothesis that sphingolipids are involved in Izh2p-dependent signaling. In this study, we demonstrate that drugs affecting sphingolipid metabolism, such as D-erythro-MAPP and myriocin, inhibit the effect of Izh2p on FET3. We also show that Izh2p causes an increase in steady-state levels of sphingoid base. Moreover, we show that Izh2p-independent increases in sphingoid bases recapitulate the effect of Izh2p on FET3. Finally, our data indicate that the Pkh1p and Pkh2p sphingoid base-sensing kinases are essential components of the Izh2p-dependent signaling pathway. In conclusion, our data indicate that Izh2p produces sphingoid bases and that these bioactive lipids probably function as the second messenger responsible for the effect of Izh2p on FET3.

Dissecting the regulation of yeast genes by the osmotin receptor
Kupchak, BR; Villa, NY; Kulemina, LV; Lyons, TJ
Biochem. Biophys. Res. Comm. 374 (2) 210-213 (2008)
<Abstract>

The Izh2p protein from Saccharomyces cerevisiae is a receptor for the plant antifungal protein, osmotin. Since Izh2p is conserved in fungi, understanding its biochemical function could inspire novel strategies for the prevention of fungal growth. However, it has been difficult to determine the exact role of Izh2p because it has pleiotropic effects on cellular biochemistry. Herein, we demonstrate that Izh2p negatively regulates functionally divergent genes through a CCCTC promoter motif. Moreover, we show that Izh2p-dependent promoters containing this motif are regulated by the Nrg1p/Nrg2p and Msn2p/Msn4p transcription factors. The fact that Izh2p can regulate gene expression through this widely dispersed element resents a reasonable explanation of its pleiotropy. The involvement of Nrg1p/Nrgp2 in Izh2p-dependent gene regulation also suggests a role for this receptor in regulating fungal differentiation in response to stimuli produced by plants. (C) 2008 Elsevier Inc. All rights reserved.

Heterologous expression of human mPR alpha, mPR beta and mPR gamma in yeast confirms their ability to function as membrane progesterone receptors
Smith, JL; Kupchak, BR; Garitaonandia, I; Hoang, LK; Maina, AS; Regalla, LM; Lyons, TJ
Steroids 73 (11) 1160-1173 (2008)
<Abstract>

The nuclear progesterone receptor (nPR) mediates many of the physiological effects of progesterone by regulating the expression of genes, however, progesterone also exerts non-transcriptional (non-genomic) effects that have been proposed to rely on a receptor that is distinct from nPR. Several members of the progestin and AdipoQ-Receptor (PAQR) family were recently identified as potential mediators of these non-genomic effects. Membranes from cells expressing these proteins, called mPR alpha, mPR beta and mPR gamma, were shown to specifically bind progesterone and have G-protein coupled receptor (GPCR) characteristics, although other studies dispute these findings. To clarify the role of these mPRs in non-genomic progesterone signaling, we established an assay for PAQR functional evaluation using heterologous expression in Saccharomyces cerevisiae. Using this assay, we demonstrate unequivocally that mPR alpha, mPR beta and mPR gamma can sense and respond to progesterone with EC50 values that are physiologically relevant. Agonist profiles also show that mPR alpha, mPR beta and mPR gamma are activated by ligands, such as 17 alpha-hydroxyprogesterone, that are known to activate non-genomic pathways but not nPR. These results strongly suggest that these receptors may indeed function as the long-sought-after membrane progesterone receptors. Additionally, we show that two uncharacterized PAQRs, PAQR6 and PAQR9, are also capable of responding to progesterone. These mPR-like PAQRs; have been renamed as mPR delta (PAQR6) and mPR epsilon (PAQR9). Additional characterization of mPR gamma and mPRa indicates that their progesterone-dependent signaling in yeast does not require heterotrimeric G-proteins, thus calling into question the characterization of the mPRs; as a novel class of G-protein coupled receptor. (c) 2008 Elsevier Inc. All rights reserved.

Probing the mechanism of FET3 repression by Izh2p overexpression
Kupchak, BR; Garitaonandia, I; Villa, NY; Mullen, MB; Weaver, MG; Regalla, LA; Kendall, EA; Lyons, TJ
Biochim. Biophys. Acta 1773 (7) 1124-1132 (2007)
<Abstract>

We previously reported a role for the IZH2 gene product in metal ion metabolism. Subsequently, Izh2p was also identified as a member of the PAQR family of receptors and, more specifically, as the receptor for the plant protein osmotin. In this report, we investigate the effect of Izh2p on iron homeostasis. We show that overproduction of Izh2p prevents the iron-dependent induction of the Fet3p component of the high-affinity iron-uptake system and is deleterious for growth in iron-limited medium. We demonstrate that the effect of Izh2p requires cAMP-dependent kinase and AMP-dependent kinase and is not mediated by general inhibition of the Aft1p iron-responsive transcriptional activator. We also show that lzh2p-overproduction negatively regulates Nrg1p/Nrg2p- and Msn2p/Msn4p-dependent reporters. Furthermore, we show that the Nrg1p/Nrg2p and Msn2p/Msn4p pairs are epistatic to each other with respect to their effects on FET3 expression. Finally, we show that the mechanism by which PAQR receptors activate signal transduction pathways is likely to be conserved from yeast to humans. (C) 2007 Elsevier B.V. All rights reserved.

Transport and Storage of Metal Ions in Biology
Lyons, TJ; Eide, DJ
Biological Inorganic Chemistry: Structure and Reactivity , ed. I. Bertini, H. Gray, E. Stiefel, and J.S. Valentine , University Science Books 57-77 (2007)

Zinc in yeast: mechanisms involved in homeostasis
Regalla, LM; Lyons, TJ
Molecular Biology of Metal Homeostasis and Detoxification: From Microbes to Man , ed. Springer , Markus J. Tamás and Enrico Martinoia 37-58 (2006)

(View all publications by Thomas Lyons)