Browsing by Author "Roman, Gregg"
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Item A Novel Function for Juvenile Hormone in Male Courtship Behavior of Drosophila melanogaster(2013-08) Wijesekera, Thilini P. 1976-; Dauwalder, Brigitte; Roman, Gregg; Williams, Cecilia M.; Mattox, WilliamJuvenile hormone is a significant insect hormone controlling development as well as reproduction, migration, and social behavior. Juvenile hormone-binding proteins of many insects are similar to Drosophila melanogaster ‘Takeout’, a protein preferentially expressed males that influences courtship behavior. This raises the possibility for a role for Juvenile hormone in male courtship behavior. This hypothesis was tested by creating flies with reduced Juvenile hormone levels and examining their mating behavior. To achieve a reduction in levels, a key enzyme of the Juvenile hormone synthesis pathway was targeted by RNA interference (RNAi). Juvenile Hormone Acid Methyl Transferase (JHAMT) catalyzes one of the last steps in the hormone’s synthesis in the Corpora allata, the organ of Juvenile hormone synthesis. A new Corpora allata-specific GAL4 driver line was created and used to direct JHAMT-RNAi to these cells. Courtship assays of the resulting males showed that RNAi against JHAMT results in a mutant courtship phenotype. Constitutive reduction as well as specific adult reduction reduced courtship. The same result was also obtained by conditional genetic ablation of the Corpora allata in adults. The courtship defects could be rescued by application of the Juvenile hormone analog Methoprene shortly before behavioral testing. Together, these results show that normal adult Juvenile hormone levels are physiologically required for normal male courtship behavior. It is unknown which protein(s) transport Juvenile hormone in Drosophila melanogaster. Since Takeout has many of the characteristics of JHBPs, we tested whether Takeout binds the hormone to act as a carrier for the hormone. No binding was observed between a Baculovirus produced Takeout protein and Juvenile hormone in an in vitro binding assay. The findings presented in this dissertation demonstrate a novel and important function for Juvenile hormone in the control of male courtship behavior in Drosophila melanogaster.Item Biased Signaling by Beta-2 Adrenergic Receptor Ligands in Murine Models of Asthma(2014-08) Thanawala, Vaidehi Jatin; Bond, Richard A.; Knoll, Brian J.; Eikenburg, Douglas C.; Roman, Gregg; Hanania, Nicola A.Asthma is a chronic inflammatory disorder of the airways that affects over 300 million people worldwide. Inhaled corticosteroids (ICS) and β2 adrenergic receptor (β2AR) agonists are the mainstay of asthma therapy. However, chronic use of ICS and β2AR agonists has been associated with adverse effects and loss of control of asthma symptoms. Clinical studies have shown the beneficial effects of chronic administration of the beta-blocker nadolol in attenuating forced expiratory volume (FEV1) in mild-asthmatics. However, not all beta-blockers are beneficial in the therapy of asthma. A clinical study published by Short and colleagues showed that chronic administration of the beta-blocker propranolol in a subset of moderate asthmatics did not improve FEV1. Similar to human studies, such discrepancy in the beneficial effects of beta-blockers has also been seen in murine models of asthma. Our previous studies have shown that certain beta-blockers with inverse agonist activity such as nadolol, ICI-118,551 and high-dose metoprolol are beneficial in attenuating the inflammation and AHR associated with the murine asthma phenotype. However, other beta-blockers that are not inverse agonists, such as alprenolol, were not as effective in attenuating the murine asthma phenotype. We have also reported that, β2AR knock out (β2AR KO) mice have an attenuated asthma phenotype, indicating the requirement of the β2AR for development of the murine asthma phenotype. Moreover, nadolol does not further attenuate the asthma phenotype in the β2AR KO mice, indicating that the beneficial effects of nadolol in the murine asthma phenotype are through its activity at the β2AR. The current project investigated the β2AR activation and the signaling pathways mediating the asthma phenotype in antigen-driven murine models. Using pharmacological and genetic models to we show that constitutive activation of the β2AR is not enough and ligand-activation of the receptor is required for development of the asthma phenotype. We used six β2AR ligands with varying signaling profiles to show the role of the non-canonical extracellular signal-regulated kinases (ERK1/2) activation pathway in development of the asthma phenotype; and suggest the subset of beta-blockers capable of shutting down β2AR-ERK1/2 signaling may have a role in the chronic management of asthma.Item Dysregulation of Cationic Channels in Chronic Kidney Diseases(2016-05) Roshanravan, Hila 1985-; Dryer, Stuart E.; Roman, Gregg; Dauwalder, Brigitte; Eriksen, Jason; Wang, YanlinAccording to the Centers for Disease Control and Prevention, one in 10 American adults, has some level of chronic kidney disease (CKD), a condition characterized by reduced kidney function over time. Although recent research has uncovered many pathways and mechanisms involved in the pathophysiology of kidney diseases, this has not yet led to development of new drugs for the treatment of patients with these conditions. In this dissertation, we introduce two potential therapeutic targets for different forms of CKD. First, we discuss gating properties of the transient receptor potential cationic-6 (TRPC6) channel, then we show dysregulation of this channel in models of focal segmental glomerulosclerosis (FSGS). In a separate chapter, we introduce another channel protein, the N-methyl-D-aspartate (NMDA) receptor, as a potential therapeutic target for treatment of diabetic nephropathy. Much of this work entailed making whole-cell recordings from highly specialized kidney cells called podocytes. This technique was used to measure TRPC6 channel activity in cells in vitro, as well as in ex vivo preparations in which podocytes are still attached to the isolated glomerular capillary. Serum samples from a variety of primary FSGS patient groups were obtained from collaborators. We used the sera to treat our cells in vitro and investigate the effect of soluble factors in the patients’ serum on the activity and expression levels of TRPC6 channels. Regarding the other target, we studied the effect of NMDA inhibitors in alleviating the development of diabetic nephropathy in vivo. I concluded that TRPC6 channels are dysregulated in FSGS. This suggested that further development of TRPC6 inhibitors might be warranted as potential therapeutic agents. We observed that diabetes caused a marked increase in the expression of renal NMDA receptors, and that sustained treatment with NMDA antagonists reduces the progression of nephropathy in two mouse models of type-1 diabetes. Consequently, it is possible that this class of drugs can be useful for reducing the progression of nephropathy.Item Identification and Characterization of Sex-Specific Transcripts in the Blood-Brain Barrier of Drosophila melanogaster(2013-12) Lama, Chamala 1979-; Dauwalder, Brigitte; Roman, Gregg; Gunaratne, Preethi H.; Beckingham, Kathleen M.In Drosophila, a male displays a series of complex stereotypic acts in courting a female. This behavior is mainly controlled by male specific transcription factors that define male neuronal circuits inside the brain. In our lab, we have shown that male factors which are secreted outside of the CNS are also required for normal mating behavior. How these endocrine factors interact with the CNS is unknown. We have evidence that the Blood-Brain Barrier (bbb) plays an important role in this communication. Specific feminization of the bbb in otherwise normal males severely reduces their courtship. This suggests that male specific factors in the bbb play an important role in mating behavior. To identify these factors, I have used several genomic screens on dissected brains and isolated bbb cells. Studies that include mRNA sequencing and microarray hybridizations have identified several male-specific candidate genes with possible novel roles in courtship. I examined and found an adult role for one of the male preferentially expressed bbb-specific candidate genes, Hr46 in the regulation of male courtship behavior in the bbb. In addition, I have identified bbb-expressed microRNAs and their possible targets. I explored a role for miR-184, the most abundant miRNA in the bbb of Drosophila in male courtship. Alterations of the miR-184 levels in the adult bbb resulted in significant reduction in courtship suggesting its importance for normal male courtship in the bbb. Conditional RNAi knockdown of sinu, a miR-184 target in the adult bbb showed significant reduction in courtship indicating a physiological requirement in the bbb. Dye injection analysis shows that the the bbb permeability of miR-184 and sinu mutants is intact. quiver, another putative miR-184 target and a gene down-regulated in the mRNA sequencing experiment, was also found to affect male courtship behavior. Taken together, these data are the first to identify sex-specific transcripts in the bbb and bbb-specific miRNAs; and to reveal a novel role for miR-184 in the bbb for male courtship behavior of Drosophila melanogaster.Item Mutants of Drosophila Melanogaster's ABC Transporter, White, Have Altered Cholesterol-Mediated Acquisition of Memory that is Moderated with 5-HT(2017-12) Myers, Jennifer Laura 1966-; Dauwalder, Brigitte; Roman, Gregg; Ziburkus, Jokubas; Wiernasz, DianeSince the discovery of Drosophila melanogaster’s white gene over 100 years ago, white has become an important genetic tool in Drosophila research. The distinct white-eyed phenotype has allowed researchers to do seminal work on heredity, gene regulation, and whole organism studies with transgenic modification. The white gene encodes an ATP binding cassette G-subfamily (ABCG) half transporter for the precursors of eye pigments and possibly biogenic amines, 5-hydroxytryptamine (5-HT) and dopamine that are critical for olfactory learning and memory in Drosophila. ABCG transporters are also known for their roles in lipid transport and homeostasis. In recent years, several behavioral differences in learning and memory have been found in white mutants that are independent of eye color and associated with 5-HT. This study characterizes a novel defect in the acquisition of olfactory memory of white mutants using the well-established classical conditioning paradigm. Classical olfactory conditioning pairs odors to shock and requires the fly to discriminate between shock paired odors and unpaired odors. The White transporter was found to be both necessary and sufficient for wild-type acquisition. 5-HTP feeding can modulate acquisition. Low 5-HT in the heads of flies, however, is not sufficient to simulate the poor acquisition seen in white mutants. This is in agreement with the normal acquisition seen in mutants of known white binding partners, scarlet1 and brown1, and high levels of 5-HT found in the heads of white1118 mutants using HPLC. Two other Drosophila ABCG mutants (CG3164MI and CG17646MI) with putative roles in cholesterol homeostasis also have poor acquisition both as homozygotes and as double heterozygotes with white1118. Mutant alleles of these ABCGs and white1118 are all partially dominant to wild-type alleles. The white1118 mutants have better acquisition with low levels of dietary cholesterol compared to high levels (0.01 mg/ml) in contrast to Canton-S wild-type flies that have better acquisition with high cholesterol feedings. Fly mass for females and males was unchanged between genotypes and cholesterol treatment. Taken together, these data suggest that the white gene has altered cholesterol homeostasis that mediates early acquisition of memory and is sensitive to 5-HT signaling.Item Report on Open Access Publishing for the Research and Scholarship Committee of the Faculty Senate with Recommendations(2016-03-03) Roman, Gregg; Fox, George E.; Bronicki, Jacqueline; Thompson, SantiIn this report, we discuss the need for the University of Houston to take an active role in managing open access (OA) publishing for the faculty and librarians.Item Sequence - Structure – Function: Analysis of the Takeout Protein Superfamily Using D. Melanogaster Courtship as a Model System(2015-12) Saurabh, Sumit 1981-; Dauwalder, Brigitte; Roman, Gregg; Zufall, Rebecca A.; Dierick, Herman A.Courtship behavior is a well characterized and extensively studied complex innate behavior in D. melanogaster adult males. Our lab has previously established the role of the secreted protein Takeout (To) in courtship behavior. To is a small soluble 23kD protein encoded by a gene that belongs to a family of 23 paralogs in D. melanogaster. To is sex-preferentially expressed in the fat body surrounding the brain in male flies. Homologs of To have been documented in other species as well. The relationship between these family members, however, remains unclear. Here, in this work I show that paralogs CG13618 and CG16820 can rescue the courtship behavior of to1 mutants. I also show that a To ortholog from A. aegypti (Diptera) and a homolog from the Light Brown Moth Epiphyas postvittana (Lepidoptera) can also rescue the courtship defects in to1 mutants. In this study I have also established the functional significance of conserved residues in motif-2 of the To protein sequence. Using site-directed mutagenesis I show that on mutating the conserved residues in motif-2 to Alanines, To only partially retains its function. Further, I also show that the hemolymph Juvenile Hormone Binding Protein (hJHBP) from Silkworm (Bombyx mori), which has very low sequence identity (31%identity, 34% sequence coverage) can partially substitute for To. I have used protein structure modeling programs to examine the sequences and structural features of the rescuing proteins and identified residues that might be critical in the ligand binding by To. Taken together my experiments establish that distant To homologs from other species and within D. melanogaster can functionally substitute for To in courtship behavior, despite their limited sequence similarities. Moreover, hJHBP from Bombyx mori can partially substitute for To suggesting that although To and hJHBPs have diverged and belong to two separate protein families they have partially retained their function. Partial rescue of courtship behavior by hJHBP also suggests that hJHBPs might be able to bind the same ligand as To.Item Serotonergic Modulation of Novelty Habituation During Exploration in Drosophila(2019-08) de la Flor, Miguel Angel 1966-; Dauwalder, Brigitte; Roman, Gregg; Dierick, Herman A.; Sater, Amy K.; Wiernasz, DianeIn a natural environment, animals must attend to and process countless streams of stimuli. Novel stimuli, or a change in the environment, may signal an opportunity for food, mates or indicate danger. A novel stimulus may elicit an approach response motivating an animal to inspect and learn about the new environmental feature. Locomotor exploration allows an animal to gain information about the features in the environment. However, animals can attend to only one stimulus at a time. Once the aim of exploring has been achieved, the exploratory behaviors triggered by novelty should cease to allow the animal to attend to other tasks. Novelty habituation is the process whereby an animal gradually decreases behaviors elicited by novelty, as the unfamiliar transitions to familiar. Herein, this study demonstrates that the decrease in locomotor activity Drosophila display in the open-filed arena is habituation to the novelty presented by the arena. In addition, experiments presented here show that serotonin signaling modulates locomotor activity, that the 5-HT1A receptor may be required in / and neurons of the mushroom bodies for locomotor modulation and that activation of the Dorsal Paired Medial neurons and possibly the Posterior Lateral Protocerebrum neurons is sufficient to decrease locomotor activity in the open-field arena. These data suggest a putative serotonergic circuit that modulates locomotor exploration in response to plasticity in the mushroom bodies as novelty transitions to familiarity.Item Sex Differences in Binge Alcohol-Induced Brain Damage and Recovery of Function(2016-05) Maynard, Mark E.; Leasure, J. Leigh; Neighbors, Clayton; Roman, Gregg; Roysam, BadrinathEvidence suggests that women are more sensitive to the neurotoxic effects of alcohol and more vulnerable to the adverse medical consequences of heavy alcohol consumption than men. Despite this few studies have directly compared the consequences of binge alcohol between the male and female brain, and the mechanisms that underlie increased female vulnerability remain poorly understood. The present studies investigated sex differences in alcohol-induced neurodegeneration, and associated cognitive deficits and disruption of trophic support, using a rodent model of an alcohol use disorder (AUD). Binge exposure resulted in a significant loss of granule neurons and significantly more degenerating and dead cells in the hippocampal dentate gyrus of females compared to males. This was associated with a binge-induced spatial reference memory deficits in the Morris water maze for females but not males. Binge-induced neurodegeneration in the female hippocampus was associated with a decrease of BDNF, TrkB, CREB, and pCREB protein expression; however only BDNF was disrupted in the hippocampus of males. Further, we investigated if exercise-driven recovery from binge-induced neurodegeneration was associated with increased trophic support. One to two weeks of voluntary exercise reversed the binge-induced reduction of dentate gyrus granule neurons in females, likely via an increase in BDNF, pCREB, and Iba1 (microglia). We conclude that the female hippocampus is more sensitive to binge-induced neurodegeneration and associated cognitive consequences than males, like due to the disruption of protective tropic support. Voluntary exercise however, can enhance endogenous recovery processes by increasing trophic support that aids in recovery.Item The Role of Pre-Synaptic Munc 13-1 in Voluntary Ethanol Consumption and Tolerance(2018-08) Wooden, Jessica I.; Leasure, J. Leigh; Roman, Gregg; Das, Joydip; Kosten, Therese A.In the United States alone, approximately 18 million people suffer from alcohol use disorders. The role of the munc 13-1 pre-synaptic protein in alcohol-related behaviors has been little-studied, despite being a known site for ethanol binding. Munc 13-1 is an active zone protein that is vital for vesicle maturation at the synapse. Mice that are heterozygous for the gene regulating munc 13-1 produce synapses that, while structurally sound, show an approximate 50% decrease in this protein. Ethanol binds munc 13-1, decreasing its functional ability and likely antagonizing glutamatergic targets. Exposure to NMDA receptor antagonists has long been known to increase hippocampal neurogenesis, and it is likely that mice with inherently deficient NMDA stimulation would show the same pro-neurogenic outcomes. The current study aimed to perform a thorough behavioral analysis of mice heterozygous for munc 13-1. Few phenotypes have been identified in this specific type of mouse, and genetically engineered mice may express very different phenotypes from mutation to mutation. This project also endeavored to assess how loss of munc 13-1 affects voluntary binge and chronic alcohol consumption. Separately, munc 13-1 heterozygotes were evaluated for changes to acute functional tolerance following an injection of a standardized dose alcohol, using an accelerating rotarod as a measure of motor coordination. The last goal of the current project was to compare levels of doublecortin (DCX)-positive cells in the dentate gyrus between alcohol-exposed and alcohol-naïve mice, and also compare DCX levels across wildtype and heterozygous mice. Taken together, these results will give future researchers a better understanding of the role that pre-synaptic protein munc 13-1 plays in alcohol consumption and tolerance.Item THE ROLE OF TEMPERATURE AND ADAPTIVE PHENOTYPIC PLASTICITY IN THE EVOLUTION OF DROSOPHILA MELANOGASTER MORPHOLOGICAL CLINES(2014-08) Russey, William Andrew 1986-; Frankino, W. Anthony; Azevedo, Ricardo B. R.; Cole, Blaine J.; Roman, Gregg; Roberts, Stephen P.Variation in morphology results in variation in ecologically relevant performances, which ultimately results in variation in fitness allowing for adaptive evolution. Task performances, such as flight ability, result from the proper scaling of and functional integration of numerous component traits. Morphological variation underlying ecologically relevant task performances can experience strong environmental effects in their expression, or phenotypic plasticity. Historically, the role phenotypic plasticity in adaptive evolution has been controversial, although it has garnered increased support in recent decades. Drosophila spp. are globally distributed and exhibit convergent morphological clines in flight morphology, and importantly, they also exhibit patterns of phenotypic plasticity consistent with these geographic patterns. In the work presented here, I examine if existing patterns of D. melanogaster flight morphology are adaptive regarding flight performance and fitness under the prediction of phenotype-environment matching, wherein the phenotype expressed in an environment enhances fitness in the predicted environment. In the work presented here, I demonstrate (i) phenotypic plasticity in D. melanogaster exhibits a pattern of adaptive phenotype-environment matching in which an induced phenotype is best-suited for flight at the temperature of development, (ii) the pattern of thermally-induced phenotypic plasticity facilitates the evolution of upwind flight performance at Cool and Warm flight temperatures, (iii) adaptive evolution by genetic accommodation is a dynamic process and the contribution of traits responding to selection vary and change over time, and finally, (iv) the adaptive pattern of phenotype-environment matching regarding flight performance is only partially realized as an increase to fitness, measured as survival in presence of predators. My dissertation work importantly demonstrates existing patterns of phenotype-environment matching in D. melanogaster, and demonstrates how this pattern facilitates adaptive evolution by genetic accommodation in a complex phenotype that exhibits natural, continuous variation.Item The Role of the Drosophila Dopamine 2-Like Receptor in the Blood-Brain Barrier for Male Courtship Behavior(2020-05) Love, Cameron Roger; Dauwalder, Brigitte; Kelleher, Erin S.; Roman, Gregg; Lekven, Arne C.; Bond, Richard A.Courtship in Drosophila Melanogaster is an extensively studied and well characterized complex innate behavior with known molecular determinants. We have previously shown that the G-protein Go-alpha and sex-specific signaling in the Blood-Brain Barrier (BBB) influence male innate courtship sustainment towards naïve females. Here we show that the glial cells that form the BBB modulate male courtship behavior through the dopamine receptor D2R. The subperineurial glia cells (SPG) form the BBB by providing a contiguous barrier, connected by septate junctions, at the interface between the hemolymph and the brain. While the neural circuits required to produce scripted actions in the fly, such as the steps for courtship, have been widely investigated, much less is known about how the circuitry and functions underlying the fly behavior is influenced by cell- non-autonomous molecular processes. We have previously shown that male-specific molecules in the BBB regulate male courtship. We identified the Dopamine-2 like receptor (D2R) RNA as one of a number of sex-specifically enriched BBB transcripts. D2R knockdown with RNAi or over-expression of D2R with a transgene in the SPG of adult males significantly reduces courtship. Knockdown or overexpression of D2R with ubiquitous neuronal drivers or SPG knockdown specifically during development has no effect on the courtship index. D2R mutant flies have courtship defects that can be rescued by expression of wildtype D2R in the BBB of adult males. D2R likely signals through Go and beta-arrestin, to maintain full male courtship levels. Our results indicate an expanded role for dopamine signaling in the glial cells that surround the brain and provide a critical time frame for its action in behavior.