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Current and Past Trainees (alphabetical listing) |
|
Erin Allen |
Molinate is a thiocarbamate herbicide, commonly used as a pre-emergent in rice patty fields, which has been shown to cause neurotoxicity. Molinate undergoes oxidation to molinate sulfoxide and is further oxidized to molinate sulfone, an electrophile reactive toward thiols. Previous work demonstrated a decrease in aldehyde dehydrogenase (ALDH) activity in rats treated with molinate. These findings suggest the sulfone metabolite of molinate, and perhaps the sulfoxide metabolite, to be an inhibitor of ALDH, potentially modifying the active-site cysteine. ALDH is an enzyme important in the catabolism of many neurotransmitters, and inhibition of these catabolic processes may lead to the accumulation of neurotoxic metabolites such as 3,4-dihydroxyphenylacetaldehyde (DOPAL). The hypothesis of my current research is molinate and both metabolites inhibit ALDH through modification of the active-site cysteine, and molinate sulfone is the most potent inhibitor. To test this hypothesis, molinate sulfoxide and molinate sulfone were synthesized, and various model systems were used, including rat brain ALDH, rat striatal mitochondria, synaptosomes, and human recombinant ALDH. The enzyme activity was monitored in each of these model systems and the relative potency of each inhibitor determined. Preliminary results demonstrate molinate, molinate sulfone, and molinate sulfoxide inhibit ALDH activity, causing an increase of the endogenous neurotoxin, DOPAL, and molinate sulfone is the most potent ALDH inhibitor. Future work will include elucidating the mechanism of ALDH inhibition for each of these inhibitors, and analyzing the effects of these inhibitors on dopaminergic PC6-3 cells. Outstanding Poster Presentation Award, Central States Society of Toxicology Meeting - 2007 |
B. Kevin Anderson |
Gene therapy has not reached its full potential in the prevention and treatment of disease. Non-viral vectors are gaining wide utility as a means of gene delivery due to the control one has when synthesizing a carrier, which can lead to increases in efficacy and safety. Non-viral vectors can be constructed efficiently and with high reproducibility. In addition, non-viral DNA condensates can be dosed repeatedly with little or no immune response or cytotoxicity. Two important elements in designing a non-viral vector are to design a compound which targets a specific receptor or cell type, and to devise a means of protecting therapeutic DNA from intracellular degradation and DNAses. The focus of my research in the Rice lab is to synthesize DNA carrier molecules with a ligand specific to the mannose receptor, and with properties which render it resistant to intracellular degradation. Because our DNA condensates are mannoseylated, we have been able to target Kupffer cells in-vivo with high specificity. In our future work we will be targeting Dendritic cells bearing the SIGN receptor, a related C-type lectin which binds mannose. DC-SIGN mediates transient adhesion with T-cells, and this contact is essential in initiating a primary immune response. |
Robert Dallapiazza | Chemical synapses are highly dynamic structures that are capable of undergoing structural and physiological changes in response to the pattern and intensity of their stimulation. These activity-dependent changes, termed synaptic plasticity, have long been recognized as a physiological substrate for learning and memory. Long-term potentiation (LTP) is the most commonly studied form of synaptic plasticity. Induction of LTP is critically dependent on the activation of N-methyl-D-aspartate-type glutamate receptors (NMDA-R) and Ca2+/Calmodulin-Dependent Kinase II (CaMKII). Postsynaptic calcium influx through NMDA-R triggers the activation of CaMKII, which translocates to the postsynaptic signaling sites through its interactions with the NMDA-R subunits NR1 and NR2B. The significance of CaMKII’s translocation is not fully known, however we hypothesize that it is an early molecular event that is necessary for the expression of synaptic plasticity. Our laboratory has developed two strains of mice with targeted mutations to NR1 and NR2B that are unable to bind to CaMKII. These two strains of mice will be used to test the significance of CaMKII translocation to the postsynaptic density. Biochemical analysis of forebrain homogenates in NR2B knock-in mice demonstrates a dramatic reduction in the association of NR2B and CaMKII compared to WT littermates. Our preliminary electrophysiological studies suggest that basal synaptic transmission is not altered in NR2B KI mice, which demonstrate similar input-output relationships and paired-pulse facilitation. However, NR2B KI mice are deficient in their ability to exhibit LTP, demonstrating only 122 ± 4% potentiation compared to WT litter-matched controls with 157 ± 7% potentiation. These results suggest that NR2B-mediated targeting of CaMKII to the postsynaptic density plays a critical role in LTP. Recipient of Kirschstein NRSA Individual Predoctoral Fellowship (2007-2009) |
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Justin Drake |
Prostate cancer is the second leading cause of death in aging men. The cause of death is not a result of the primary tumor growth, but rather complications due to metastatic tumors. Prostate cancer predominantly metastasizes to bone inducing osteoblastic lesions. A potent vasoconstrictor, endothelin-1 (ET-1), has recently been implicated in prostate cancer bone metastasis. However, the mechanism in which tumor-derived ET-1 interacts in the bone microenvironment is not understood. To this end, I am studying the role of ET-1 in prostate cancer bone metastasis. I have engineered a luciferase expressing prostate cancer cell line, 22Rv1, that will be introduced into SCID mice using intracardiac injections. The technique of bioluminescent imaging (BLI) will be used to monitor tumor growth and colonization patterns of 22Rv1-luciferase cells in vivo. I plan to modulate tumor-derived ET-1 by RNAi and ET-1 specific drug inhibitors in vivo. Using this model, I plan to investigate the cellular and molecular biology pertaining to prostate cancer metastasis to bone. Winner: 2006 Graduate Student
Poster Session - Carver College of Medicine |
Amanda Fenner |
Heparan sulfate (HS) is a highly-charged, polysulfonated, polysaccharide located on the surface and in the extracellular matrix of mammalian cells. HS plays a profound role in the recognition of, adhesion to, and/or infectivity of host cells by many pathogenic organisms including plasmodia, yersina pestis, listeria monocytogenes, and giardia lamblia and in the progression of non-pathogenic diseases such as cancer, Alzheimer’s, and atherosclerosis. Molecules that selectively bind HS-binding proteins and block HS-protein interactions involved in host-pathogen interactions are needed for the development of therapeutic agents to block invasion, replication and/or virulence of these diseases. The Kerns lab has recently identified chemically-modified heparin derivatives that selectively bind and block HS-binding proteins. My research is focused on the design and synthesis of N-acylated/O-sulfonated derivatives of naturally occurring amino sugars as novel, selective inhibitors of HS-protein interactions. I will also employ enzymatic depolymerization of naturally occurring polysaccharides to create size-defined oligosaccharides for further chemical modification as inhibitors of HS-binding proteins. HS-protein binding studies will be performed to evaluate the modified saccharides as potential inhibitors of host-pathogen interactions as well inhibitors of heparanase and fibroblast growth factors, which are targets for cancer treatment. Ultimately, my goal is to employ my chemically modified and/or biocatalytically-derived saccharides that selectively bind bacterial surface proteins as chemical probes to study HS binding versus HS bridging mechanisms in bacteria-host cell interactions. |
Lawrence Gray |
My research in Dr. Khademi’s laboratory focuses on the structure and function of biologically important prokaryotic membrane proteins. About 30% of the genes in a cell encode membrane proteins. However, less than 0.5% of entries in PDB (a database for the coordinates of proteins with known structures) are membrane proteins. While more than 60% of drug receptors are membrane proteins, our knowledge of their structures and functions are very limited. My research goal is to determine the x-ray structure of essential membrane proteins from H. pylori. I will also conduct functional assays to determine the mechanism of action of these membrane proteins. |
Melissa Hall |
One of the primary focuses of Dr. Amnon Kohen's lab is the study of chemical catalysis in biological systems. My research focused on resolving the catalytic cascade, using pre-steady state kinetics, of flavin-dependent thymidylate synthase (FDTS) isolated from Thermotoga maritime (tmFDTS). In general, thymidylate synthase (TS) activity is essential for any living organism because it catalyzes the last committed step in the biosynthesis of thymidine. Preliminary studies have indicated that the FDTS reaction is substantially different than that of classical TS (which is found in humans). These mechanistic differences suggest that resolution of the catalytic cascade (the order of substrate binding and product release, formation of intermediate complexes, etc.) will contribute to the design and synthesis of effective inhibitors. These inhibitors will be tested in classical TS, as a measure of possible toxicity. Promising compounds will then be examined as in vitro and in vivo antibiotic drugs. |
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Amy Halt |
The post-synaptic density (PSD),
a thickened area of the post-synaptic membrane visible under
electron microscope, contains scaffolding proteins that cluster
glutamate receptors at the synapse. N-methyl-D-aspartate (NMDA)
receptors are concentrated along with a-amino-3-hydroxy-5-methyl-4-isoxazolepropionate
(AMPA) receptors within the PSD. Ca2+ influx through the NMDA
receptor plays a central role in mediating cellular changes
leading to increased synaptic strength. Ca2+ influx through
the NMDA receptor following excessive glutamate release is also
thought to underlie many of the detrimental signaling cascades
leading to cell death following cerebral ischemia. Ca2+/calmodulin-dependent
protein kinase II (CaMKII), a serine/threonine kinase, is important
in mediating cellular responses to Ca2+ including changes subsequent
to both LTP and ischemia. Activity-dependent subcellular localization
of CaMKII within the PSD is mediated through its binding to
the NMDA receptor. CaMKII strongly binds to two subunits, NR1
and NR2B. Two regions within the C-terminal cytosolic domain
of NR2B, residues 839-1120 (NR2B-P) and residues 1290-1309 (NR2B-C),
bind CaMKII. Autophosphorylation of CaMKII is required for binding
at NR2B-P, whereas the presence of Ca2+/calmodulin is sufficient
to induce CaMKII binding at NR2B-C. The C-terminal cytosolic
portion of NR1 contains 4 regions, C0, C1, C2 and C2’
that are differentially present in 4 isoforms. Autophosphorylated
CaMKII binds to the 30 residue-long, membrane-proximal, C0 region
of NR1. The binding domain for CaMKII is located at the C-terminal
end of the region and covers residues 847-858 where it competes
for binding with Ca2+/calmodulin and a-actinin-2. Functional
roles for CaMKII binding to the NMDA receptor could include
targeting of CaMKII to the PSD, regulation of NMDA receptor
activity, targeting of CaMKII to substrates for phosphorylation,
and structural modification of the PSD. The goal of my research
is to further clarify the functional significance of CaMKII
binding to the NMDA receptor. My research utilizes knock-in
mice with mutations in the CaMKII binding domains on NR1 and
NR2B. Receipient of Kirschstein NRSA Individual Predoctoral Fellowship (2006-2008) |
Beixin (Julie) He |
Work in the Anderson laboratory focuses around the multifunctional enzyme calcium/calmodulin dependent protein kinase II (CaMKII) and specifically its role and activation in the hypertropic heart. Previous work in the lab has demonstrated that CaMKII is a downstream signal in the beta-adrenergic receptor cascade. Interestingly, beta-adrenergic signaling activates myocyte enhancer factor 2 (MEF2), a transcription factor implicated in cardiac hypertrophy. Literature shows that this activation is dependent on the derepression of histone deactylases (HDACs) via CaMKII phosphorylation and subsequent nuclear exportation of HDAC. Alternatively, my project proposes that HDAC removal is primarily through its interaction with calmodulin in a competitive manner that reflects the surge in intracellular calcium and calmodulin activation that accompanies CaMKII signaling. My lab is equipped with mouse models of membrane (MEM) and nuclear-targeted (NLS) CaMKII inhibition by myocardial-restricted over-expression of a highly selective inhibitory peptide. With these tools and selectively mutated HDAC constructs, I aim to separately investigate the role of CaMKII at membrane delimited calcium homeostatic proteins and in the nucleus, a distinction that is important to the activation of MEF2. |
Patrick Houlihan |
Mitochondria are dynamic organelles that rapidly and reversibly undergo fission and fusion (termed mitochondrial fission and fusion, or MFF). Both of these processes reflect the developmental and situational state of the cell and require a number of evolutionarily conserved proteins. Data from Stefan Strack’s lab demonstrate that reversible phosphorylation at the outer mitochondrial membrane (OMM) regulates mitochondrial shape in a bi-directional manner: phosphorylation by protein kinase A (PKA) promotes mitochondrial fusion, whereas dephosphorylation by protein phosphatase 2A (PP2A) shifts MFF equilibrium toward fission by way of the Bβ2 PP2A regulatory subunit. One proposed effect of these alterations in organelle morphology is the shaping of calcium signals. In addition to performing cellular respiration and acting as signaling platforms, mitochondria efficiently buffer and store cytosolic calcium. Accordingly, shifts in MFF equilibrium appear to be essential for controlling such Ca2+-dependent functions as synaptic plasticity and neuronal survival during/following a cytotoxic insult (stroke). My preliminary data and other data from Yuriy Usachev’s lab suggest that neuronal Ca2+ signaling is strongly affected by changes in mitochondrial architecture. The mechanisms underlying these changes, as well as pharmacological manipulation of these dynamic events are my main research interests. |
Uche Maduka |
Chronic pain caused by various diseases affects 50 million people within the US alone, probably billions of people worldwide, but yet, mechanisms of chronic pain are still poorly understood. Long term potentiation (LTP) is a process by which synapses can change in transmission strength as a result of high frequency stimulation. Previous research has shown that LTP is dependent on calcium influx through the NMDA receptor (NMDAR) as well as activation of the Ca2+ / Calmodulin-dependent protein kinase II and its binding to subunits of the NMDA receptor. Earlier work in the laboratory of Dr. Hell has defined the binding sites for CaMKII on the NR1 and NR2B subunits of the NMDA receptor. We have also found that interfering with CaMKII binding to the NR1 and NR2B subunits inhibits hippocampal LTP and memory. Recently, researchers have begun to appreciate the parallels between learning and chronic pain, as nerve injuries can lead to molecular changes within the spinal cord through processes very similar to LTP. It is our hypothesis that molecular and pharmacological ways that interfere with LTP will also inhibit the development of chronic pain. To test this hypothesis, we have developed mice with impaired CaMKII binding to the NR2B subunit of the NMDAR. Protein levels of CaMKII, NMDAR subunits and other postsynaptic proteins in these mice are comparable to those in wild type mice. These mice are therefore a great tool with which we can investigate the role of the CaMKII and NMDAR association in chronic pain. If we find that impaired CaMKII association with the NMDAR interferes with chronic pain, then we will move forward with the development of organic agents that interfere with CaMKII binding to the NMDAR as novel therapeutics for the treatment of chronic pain. |
Nolan Mente |
The focus of my research is the synthesis of the schweinfurthins and related natural products (e.g. vedelianin) that have potential as anticancer agents. Both enantiomers of 3-deoxy-5-methoxyvedelianin have been synthesized and determined to have significant cytotoxicity in the NCI’s 60 cell line assay (0.41 µM and 0.13 µM, respectively). Interestingly, the activity profile of the R,R,R-enantiomer does not match that of the S,S,S-enantiomer, leaving open the possibility that the two enantiomers work at different targets and/or by a different mechanism of action. Work continues on the development of analogs designed to increase the potency and illuminate what is believed to be a novel mechanism of cytotoxicity. University of Iowa Presidential Scholar |
|
Garrett Rettig |
The focus of the Rice lab is to design novel peptide-DNA conjugates to
mediate non-viral gene delivery and gene expression in vivo. A major barrier
in non-viral gene delivery is targeting plasmid DNA (pDNA) to the nucleus. The
T antigen of SV40 is widely known to possess a short, lysine-rich peptide
that is sufficient to mediate nuclear localization. Therefore, we are pursuing
methods to incorporate a peptide containing a known nuclear localizing sequence
(NLS) into our gene formulations. Recipient of AFPE Pre-Doctoral Fellowship (2005-2006) |
Tyson Shepherd |
T-cell lymphoma Invasion and Metastasis 1 (Tiam1) protein is a Guanine Exchange Factor (GEF) that specifically activates the Rho-family GTPase Rac1. In mammalian cells, spacial and temporal regulation is achieved via several distinct protein-protein interaction domains. The Fuentes Lab applies biophysical techniques to understand how each of these domains contribute to Tiaim1 activity. Included in this set of domains is a Ras Binding (RB) domain just N-terminal to a Post-synaptic density-95/Discs large/Zonula occludens-1 (PDZ) domain. Tiam1 GEF activity is thought to increase upon activated Ras binding to the RB domain, while the functional importance of the PDZ domain remains a mystery. My research is directed at understanding the structural and functional relationship between these two domains both alone and in the context of the full length Tiam1. To understand these domains further, we express each domain separately and together with various surrounding domains. The structure of the PDZ domain was solved in both free form and bound to model peptide ligand. Nuclear magnetic resonance was used to check the binding affinity of the ligand and to investigate intermolecular dynamics upon peptide binding. Further structure and function work will be carried out on neighboring domains in the Tiam1 protein. We are also examining the effect of mutations in the binding pocket of the PDZ domain contained within the full-length Tiam1 in vivo. |
Jacqueline Smits
|
Farnesylation of Ras proteins is a critically important step in post-translational processing which is required for the membrane association of Ras proteins. Ras proteins cycle between a GTP bound active and a GDP bound inactive state which serve as signaling molecules and regulate cell proliferation. The ability to track Ras proteins in vivo will provide information about the intracellular role of Ras and the processes that occur subsequent to the post-translational modification of these proteins. The ability to affect cell proliferation makes the Ras family proteins and the process of farnesylation interesting targets for studies aimed at the development of new anti-cancer agents. |
Michelle Thein |
Evidence suggests that mental illness may be related to both peripheral and central inflammatory processes. Pro-inflammatory cytokines are small signaling molecules released during infection or inflammation. Notably, these molecules are released in response to acute myocardial infarction. Major depressive disorder has been found to be increasingly associated with myocardial infarction. Furthermore, the development of major depressive disorder has been found to impact on morbidity and mortality following myocardial infarction. As serotonin has long been understood to play a role in major depressive disorder as well as other forms of mental illness, I am investigating the relationship between the pro-inflammatory cytokines and serotonin, currently focusing on TNF-α and serotonin synthesis. |
|
Kevin Tidgewell |
The Prisinzano Lab investigates the interactions of drugs with the central nervous system (CNS). Through the elucidation of the mechanism of action of these drugs, we can begin to more clearly understand the underlying mechanism of addiction as well as begin to develop novel therapeutics for its treatment. In particular, we are very interested in the mechanism and modulation of neuropathic and chronic pain. In order to understand the mechanism of pain transmission and the pathways involved, selective agonists and antagonists are required for receptors involved in these pathways. Many pharmacological tools have come from natural sources and so the investigation of a novel natural source for opioid ligands provides a unique source for the development of new pharmacological treatments for pain. My research focuses on the novel kappa opioid ligand salvinorin A isolated from the Mexican sage Salvia divinorum. By systematically modifying salvinorin A, we will generate a better understanding of the required pharmacophore for affinity and activity at opioid receptors. Additionally, compounds which show promise (i.e. high selectivity and affinity) will be tested in in vivo pharmacological tests of nociception and drug self-administration. This plan of investigation will follow the complete process of drug discovery from isolation, to chemical synthesis and pharmacological testing in vivo. Recipient of National Medicinal Chemistry Symposium Travel Award (2006) |
Arianne Waseen |
Myelin
oligodendrocyte protein (MOG) is a protein found exclusively in
the myelin wrapping cells of the central nervous system, oligodendrocytes.
The oligodendrocytes function to insulate neurons with several
tightly packed lipid-enriched layers. MOG’s function is
unknown although some research suggests that it may be a receptor.
In most mammals, there is only one variant of MOG, but primates
express seven different splice variants The cytoplasmic tail of mouse MOG was used in a yeast-two-hybrid assay to identify putative interacting partners. A member of the stathmin family was identified as an interacting partner. Stathmin and stathmin-like proteins act as microtubulin destabilizing proteins but it is unclear which member of the stathmin family interacts with MOG. In the future I plan on determining which stathmin member interacts with MOG, and also characterizing this interaction. Since the four amino acids in MOG critical for interaction with stathmin are not found in five of the seven MOG splice variants, it is likely that these other variants have different interacting partners. To identify these partners, yeast-two-hybrid screens are currently being performed using peptides specific to two of the MOG splice variants. |
supported by the National Institute of General Medical Sciences.
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