Life Sciences Commons^™

Glia Excitation In The Cns Modulates Intact Behaviors And Sensory-Cns-Motor Circuitry, Shelby Mccubbin, Douglas A. Harrison, Robin L. Cooper

Biology Faculty Publications

Glial cells play a role in many important processes, though the mechanisms through which they affect neighboring cells are not fully known. Insights may be gained by selectively activating glial cell populations in intact organisms utilizing the activatable channel proteins channel rhodopsin (ChR2XXL) and TRPA1. Here, the impacts of the glial-specific expression of these channels were examined in both larval and adult Drosophila. The Glia > ChR2XXL adults and larvae became immobile when exposed to blue light and TRPA1-expressed Drosophila upon heat exposure. The chloride pump expression in glia > eNpHR animals showed no observable differences in adults or larvae. In …

The Effect Of Optogenetically Activating Glia On Neuronal Function, Cecilia Pankau, Shelby Mccubbin, Robin L. Cooper

Biology Faculty Publications

Glia, or glial cells, are considered a vital component of the nervous system, serving as an electrical insulator and a protective barrier from the interstitial (extracellular) media. Certain glial cells (i.e., astrocytes, microglia, and oligodendrocytes) within the CNS have been shown to directly affect neural functions, but these properties are challenging to study due to the difficulty involved with selectively-activating specific glia. To overcome this hurdle, we selectively expressed light-sensitive ion channels (i.e., channel rhodopsin, ChR2-XXL) in glia of larvae and adult Drosophila melanogaster. Upon activation of ChR2, both adults and larvae showed a rapid contracture of body wall …

Neurotensin Regulates Proliferation And Stem Cell Function In The Small Intestine In A Nutrient-Dependent Manner, Stephanie A. Rock, Kai Jiang, Yuanyuan Wu, Yajuan Liu, Jing Li, Heidi L. Weiss, Chi Wang, Jianhang Jia, Tianyan Gao, B. Mark Evers

Surgery Faculty Publications

BACKGROUND & AIMS: Intestinal stem cells (ISCs) are sensitive to dietary alterations and nutrient availability. Neurotensin (NT), a gut peptide localized predominantly to the small bowel and released by fat ingestion, stimulates the growth of intestinal mucosa under basal conditions and during periods of nutrient deprivation, suggesting a possible role for NT on ISC function.

METHODS: Leucine-rich repeat-containing G-protein coupled receptor 5-Enhanced Green Fluorescent Protein (Lgr5-EGFP) NT wild type (Nt^+/+) and Lgr5-EGFP NT knockout (Nt^-/-) mice were fed ad libitum or fasted for 48 hours. Small intestine tissue and crypts were examined by gene …

Highly Contiguous Assemblies Of 101 Drosophilid Genomes, Bernard Y. Kim, Jeremy R. Wang, Danny E. Miller, Olga Barmina, Emily Delaney, Ammon Thompson, Aaron A. Comeault, David Peede, Emmanuel R. R. D'Agostino, Julianne Pelaez, Jessica M. Aguilar, Diler Haji, Teruyuki Matsunaga, Ellie E. Armstrong, Molly Zych, Yoshitaka Ogawa, Marina Stamenković-Radak, Mihailo Jelić, Marija Savić Veselinović, Marija Tanasković, Jeremy S. Davis

Biology Faculty Publications

Over 100 years of studies in Drosophila melanogaster and related species in the genus Drosophila have facilitated key discoveries in genetics, genomics, and evolution. While high-quality genome assemblies exist for several species in this group, they only encompass a small fraction of the genus. Recent advances in long-read sequencing allow high-quality genome assemblies for tens or even hundreds of species to be efficiently generated. Here, we utilize Oxford Nanopore sequencing to build an open community resource of genome assemblies for 101 lines of 93 drosophilid species encompassing 14 species groups and 35 sub-groups. The genomes are highly contiguous and complete, …

Editorial: Invertebrate Neuroscience: Contributions From Model And Non-Model Species, Maria P. Fernandez, Clare C. Rittschof, Jimena A. Sierralta

Entomology Faculty Publications

No abstract provided.

Effect Of Temperature On Heart Rate For Phaenicia Sericata And Drosophila Melanogaster With Altered Expression Of The Trpa1 Receptors, Nicole T. Marguerite, Jate Bernard, Douglas A. Harrison, David Harris, Robin L. Cooper

Biology Faculty Publications

The transient receptor potential (TrpA—ankyrin) receptor has been linked to pathological conditions in cardiac function in mammals. To better understand the function of the TrpA1 in regulation of the heart, a Drosophila melanogaster model was used to express TrpA1 in heart and body wall muscles. Heartbeat of in intact larvae as well as hearts in situ, devoid of hormonal and neural input, indicate that strong over-expression of TrpA1 in larvae at 30 or 37 °C stopped the heart from beating, but in a diastolic state. Cardiac function recovered upon cooling after short exposure to high temperature. Parental control larvae (UAS-TrpA1) …

The Roles Of Polar Cell Extensions In Drosophila Micropyle Formation, Bradford Hull

Theses and Dissertations--Biology

The Drosophila micropyle is a conserved formation utilized to allow sperm passage past the robust eggshell structure for fertilization. Micropyle formation follows a unique acellular tubulogenesis method where it is secreted and shaped by specialized follicle cells including the border cells and polar cells. In late oogenesis, the polar cells form extensions that are necessary to create the micropyle pore through which sperm enters. Previous work established that polar cell extension presence is required for micropyle pore formation. We investigated temporal requirements of extensions throughout chorion deposition and found extensions are required during the beginning and middle of choriogenesis, but …

The Effect Of Ph On Synaptic Transmission At The Neuromuscular Junction In Drosophila Melanogaster, Catherine Elizabeth Stanley

Theses and Dissertations--Biology

Synaptic transmission is the main avenue of neuronal communication and can be affected by a multitude of factors, both intracellularly and extracellularly. The effects of pH changes on synaptic transmission have been studied for many years across many different models. Intracellular acidification at the presynaptic terminal is known to occur with increased neuronal activity and can also occur in pathological conditions. The effects of these pH alterations are therefore an important area of study. Here, intracellular acidification using either propionic acid or the ammonium chloride pre-pulse technique was examined for the effects on both spontaneous and evoked synaptic transmission at …

The Effects Of A Bacterial Endotoxin On Behavior And Sensory-Cns-Motor Circuits In Drosophila Melanogaster, Oscar Istas, Abigail Greenhalgh, Robin L. Cooper

Biology Faculty Publications

The effect of bacterial sepsis on animal behavior and physiology is complex due to direct and indirect actions. The most common form of bacterial sepsis in humans is from gram-negative bacterial strains. The endotoxin (lipopolysaccharide, LPS) and/or associated peptidoglycans from the bacteria are the key agents to induce an immune response, which then produces a cascade of immunological consequences. However, there are direct actions of LPS and associated peptidoglycans on cells which are commonly overlooked. This study showed behavioral and neural changes in larval Drosophila fed commercially obtained LPS from Serratia marcescens. Locomotor behavior was not altered, but feeding …

The Effects Of A Ketone Body On Synaptic Transmission, Alexandra Elizabeth Stanback

Theses and Dissertations--Biology

The ketogenic diet is commonly used to control epilepsy, especially in cases when medications cannot. The diet typically consists of high fat, low carb, and adequate protein and produces a metabolite called acetoacetate. Seizure activity is characterized by glutamate excitotoxicity and therefore glutamate regulation is a point of research for control of these disorders. Acetoacetate is heavily implicated as the primary molecule responsible for decreasing glutamate in the synapse; it is believed that acetoacetate interferes with the transport of glutamate into the synaptic vesicles. The effects on synaptic transmission at glutamatergic synapses was studied in relation to the ketogenic diet …

Jak/Stat Signaling Regulates Gametogenesis And Age-Related Reproductive Maintenance, Michelle Suzanne Giedt

Theses and Dissertations--Biology

Cell signaling is central to integration of internal and external cues that regulate development and homeostasis. Most development is thought of as pre-adult, but limited developmental processes occur in adults. Gametogenesis incorporates elements of both these facets, with a distinct developmental plan for gamete synthesis which is regulated by integration of homeostatic inputs such as nutrient status, and environmental cues. Signaling pathways integrate and transduce information from these cues to evoke a response. A decline in homeostasis and subsequent cues occurs over time, in the case of reproductive tissues leading to a progressive loss of fertility. The Janus Kinase and …

Sumo Regulates The Activity Of Smoothened And Costal-2 In Drosophila Hedgehog Signaling, Jie Zhang, Yajuan Liu, Kai Jiang, Jianhang Jia

Markey Cancer Center Faculty Publications

In Hedgehog (Hh) signaling, the GPCR-family protein Smoothened (Smo) acts as a signal transducer that is regulated by phosphorylation and ubiquitination, which ultimately change the cell surface accumulation of Smo. However, it is not clear whether Smo is regulated by other post-translational modifications, such as sumoylation. Here, we demonstrate that knockdown of the small ubiquitin-related modifier (SUMO) pathway components Ubc9 (a SUMO-conjugating enzyme E2), PIAS (a SUMO-protein ligase E3), and Smt3 (the SUMO isoform in Drosophila) by RNAi prevents Smo accumulation and alters Smo activity in the wing. We further show that Hh-induced-sumoylation stabilizes Smo, whereas desumoylation by Ulp1 …

Population Dynamics Based On Resource Availability & Founding Effects: Live & Computational Models, Samuel Potter, Rebecca M. Krall, Susan Mayo, Diane Johnson, Kimberly J. Zeidler-Watters, Robin L. Cooper

Biology Faculty Publications

With the looming global population crisis, it is more important now than ever that students understand what factors influence population dynamics. We present three learning modules with authentic, student-centered investigations that explore rates of population growth and the importance of resources. These interdisciplinary modules integrate biology, mathematics, and computer-literacy concepts aligned with the Next Generation Science Standards. The activities are appropriate for middle and high school science classes and for introductory college-level biology courses. The modules incorporate experimentation, data collection and analysis, drawing conclusions, and application of studied principles to explore factors affecting population dynamics in fruit flies. The variables …

Pi(4)P Promotes Phosphorylation And Conformational Change Of Smoothened Through Interaction With Its C-Terminal Tail, Kai Jiang, Yajuan Liu, Junkai Fan, Jie Zhang, Xiang-An Li, B. Mark Evers, Haining Zhu, Jianhang Jia

Markey Cancer Center Faculty Publications

In Hedgehog (Hh) signaling, binding of Hh to the Patched-Interference Hh (Ptc-Ihog) receptor complex relieves Ptc inhibition on Smoothened (Smo). A longstanding question is how Ptc inhibits Smo and how such inhibition is relieved by Hh stimulation. In this study, we found that Hh elevates production of phosphatidylinositol 4-phosphate (PI(4)P). Increased levels of PI(4)P promote, whereas decreased levels of PI(4)P inhibit, Hh signaling activity. We further found that PI(4)P directly binds Smo through an arginine motif, which then triggers Smo phosphorylation and activation. Moreover, we identified the pleckstrin homology (PH) domain of G protein-coupled receptor kinase 2 (Gprk2) as an …

Optogenetic Stimulation Of Drosophila Heart Rate At Different Temperatures And Ca2+ Concentrations, Yuechen Zhu, Henry Uradu, Zana R. Majeed, Robin L. Cooper

Biology Faculty Publications

Optogenetics is a revolutionary technique that enables noninvasive activation of electrically excitable cells. In mammals, heart rate has traditionally been modulated with pharmacological agents or direct stimulation of cardiac tissue with electrodes. However, implanted wires have been known to cause physical damage and damage from electrical currents. Here, we describe a proof of concept to optically drive cardiac function in a model organism, Drosophila melanogaster. We expressed the light sensitive channelrhodopsin protein ChR2.XXL in larval Drosophila hearts and examined light‐induced activation of cardiac tissue. After demonstrating optical stimulation of larval heart rate, the approach was tested at low temperature …

Lights And Larvae: Using Optogenetics To Teach Recombinant Dna And Neurobiology, John Titlow, Heidi Anderson, Robin L. Cooper

Biology Faculty Publications

Switching genes between organisms and controlling an animal’s brain using lasers may seem like science fiction, but with advancements in a technique called optogenetics, such experiments are now common in neuroscience research. Optogenetics combines recombinant DNA technology with a controlled light source to help researchers address biomedical questions in the life sciences. The technique has gained the most traction in neurobiology—the biology of the nervous system—where specific wavelengths of light are used to control or measure the activity of neurons in transgenic organisms (i.e., those with artificially inserted genes).

These optical recording and stimulation techniques are used in nervous system …

The Jak/Stat Pathway Is Reutilized In Drosophila Spermatogenesis, Lingfeng Tang

Theses and Dissertations--Biology

In the Drosophila testis, sperm are derived from germline stem cells (GSCs) which undergo a stereotyped pattern of divisions and differentiation. The somatic cells at the tip of the testis form the hub, which is the niche for both the somatic cyst stem cells (CySCs) and GSCs. The hub expresses Upd, a ligand for the JAK/STAT pathway that has roles in the maintenance of CySCs and GSCs. Male mutants of upd3, another ligand of the JAK/STAT pathway, become sterile much earlier than the wild-type, leading to the hypothesis that similar to upd, upd3 also promotes the self-renewal of …

Dreadds In Drosophila: A Pharmacogenetic Approach For Controlling Behavior, Neuronal Signaling, And Physiology In The Fly, Jaime Becnel, Oralee Johnson, Zana R Majeed, Vi Tran, Bangning Yu, Bryan L. Roth, Robin L. Cooper, Edmund K. Kerut, Charles D. Nichols

Biology Faculty Publications

We have translated a powerful genetic tool, designer receptors exclusively activated by designer drugs (DREADDs), from mammalian systems to Drosophila melanogaster to selectively, rapidly, reversibly, and dose-dependently control behaviors and physiological processes in the fly. DREADDs are muscarinic acetylcholine G protein-coupled receptors evolved for loss of affinity to acetylcholine and for the ability to be fully activated by an otherwise biologically inert chemical, clozapine-N-oxide. We demonstrate its ability to control a variety of behaviors and processes in larvae and adults, including heart rate, sensory processing, diurnal behavior, learning and memory, and courtship. The advantages of this particular technology include the …

Usp8 Promotes Smoothened Signaling By Preventing Its Ubiquitination And Changing Its Subcellular Localization, Ruohan Xia, Hongge Jia, Junkai Fan, Yajuan Liu, Jianhang Jia

Molecular and Cellular Biochemistry Faculty Publications

The seven transmembrane protein Smoothened (Smo) is a critical component of the Hedgehog (Hh) signaling pathway and is regulated by phosphorylation, dimerization, and cell-surface accumulation upon Hh stimulation. However, it is not clear how Hh regulates Smo accumulation on the cell surface or how Hh regulates the intracellular trafficking of Smo. In addition, little is known about whether ubiquitination is involved in Smo regulation. In this study, we demonstrate that Smo is multi-monoubiquitinated and that Smo ubiquitination is inhibited by Hh and by phosphorylation. Using an in vivo RNAi screen, we identified ubiquitin-specific protease 8 (USP8) as a deubiquitinase that …

An Evolutionarily Conserved Rit Gtpase-P38 Mapk Signaling Pathway Mediates Oxidative Stress Resistance, Weikang Cai, Jennifer L. Rudolph, Susan M. W. Harrison, Ling Jin, Aubrey L. Frantz, Douglas A. Harrison, Douglas A. Andres

Molecular and Cellular Biochemistry Faculty Publications

Ras-related small GTP-binding proteins control a wide range of cellular processes by regulating a variety of effector pathways, including prominent roles in the control of mitogen-activated protein kinase (MAPK) cascades. Although the regulatory role(s) for many Ras family GTPases are well established, the physiological function for the Rit/Rin subfamily has been lacking. Here, using both knockout mice and Drosophila models, we demonstrate an evolutionarily conserved role for Rit subfamily GTPases (mammalian Rit and Rin, and the Drosophila RIC homologue) in governing survival in response to oxidative stress. Primary embryonic fibroblasts derived from Rit knockout mice display increased apoptosis and selective …

Sonic Hedgehog Dependent Phosphorylation By Ck1Α And Grk2 Is Required For Ciliary Accumulation And Activation Of Smoothened, Yongbin Chen, Noriaki Sasai, Guoqiang Ma, Tao Yue, Jianhang Jia, James Briscoe, Jin Jiang

Markey Cancer Center Faculty Publications

Hedgehog (Hh) signaling regulates embryonic development and adult tissue homeostasis through the GPCR-like protein Smoothened (Smo), but how vertebrate Smo is activated remains poorly understood. In Drosophila, Hh dependent phosphorylation activates Smo. Whether this is also the case in vertebrates is unclear, owing to the marked sequence divergence between vertebrate and Drosophila Smo (dSmo) and the involvement of primary cilia in vertebrate Hh signaling. Here we demonstrate that mammalian Smo (mSmo) is activated through multi-site phosphorylation of its carboxyl-terminal tail by CK1α and GRK2. Phosphorylation of mSmo induces its active conformation and simultaneously promotes its ciliary accumulation. We demonstrate that …

Cooperative And Antagonistic Contributions Of Two Heterochromatin Proteins To Transcriptional Regulation Of The Drosophila Sex Determination Decision, Hui Li, Janel Rodriguez, Youngdong Yoo, Momin Mohammed Shareef, Ramakrishna Badugu, Jamila I. Horabin, Rebecca Kellum

Biology Faculty Publications

Eukaryotic nuclei contain regions of differentially staining chromatin (heterochromatin), which remain condensed throughout the cell cycle and are largely transcriptionally silent. RNAi knockdown of the highly conserved heterochromatin protein HP1 in Drosophila was previously shown to preferentially reduce male viability. Here we report a similar phenotype for the telomeric partner of HP1, HOAP, and roles for both proteins in regulating the Drosophila sex determination pathway. Specifically, these proteins regulate the critical decision in this pathway, firing of the establishment promoter of the masterswitch gene, Sex-lethal (Sxl). Female-specific activation of this promoter, Sxl(Pe), is essential to females, as it provides SXL …

Analysis Of The Crmp Gene In Drosophila: Determining The Regulatory Role Of Crmp In Signaling And Behavior, Deanna Hardt Morris

University of Kentucky Doctoral Dissertations

The mammalian genome encodes five collapsin response mediator protein (CRMP) isoforms. Cell culture studies have shown that the CRMPs mediate growth cone dynamics and neuron polarity through associations with a variety of signal transduction components and cytoskeletal elements. CRMP is also a member of a protein family including the presumably ancestral dihydropyrimidinase (DHP) protein that catalyzes the second step in pyrimidine degradation. In Drosophila, CRMP and DHP proteins are produced by alternatively spliced transcripts of the CRMP gene. The alternative protein forms have a 91% sequence identity, but unique expression patterns. CRMP is found exclusively in neuronal tissues and …

Genome-Wide Analysis Of The Interaction Between The Endosymbiotic Bacterium Wolbachia And Its Drosophila Host, Zhiyong Xi, Laurent Gavotte, Yan Xie, Stephen L. Dobson

Entomology Faculty Publications

BACKGROUND: Intracellular Wolbachia bacteria are obligate, maternally-inherited, endosymbionts found frequently in insects and other invertebrates. The success of Wolbachia can be attributed in part to an ability to alter host reproduction via mechanisms including cytoplasmic incompatibility (CI), parthenogenesis, feminization and male killing. Despite substantial scientific effort, the molecular mechanisms underlying the Wolbachia/host interaction are unknown.

RESULTS: Here, an in vitro Wolbachia infection was generated in the Drosophila S2 cell line, and transcription profiles of infected and uninfected cells were compared by microarray. Differentially-expressed patterns related to reproduction, immune response and heat stress response are observed, including multiple genes that have …

The Influence Of Ca2+ Regulation In Synaptic Facilitation Of Motor Nerve Terminals In Crayfish And Drosophila As Well As In The Physiological Regulation Of Larval Drosophila Heart, Mohati Desai-Shah

University of Kentucky Doctoral Dissertations

Intracellular Ca²⁺ ions are highly regulated in animal cells for them to function normally. Since the tight regulation of [Ca²⁺]i is so ubiquitous among cells, it is not surprising that altered function in [Ca²⁺]i regulation is associated with a myriad of disease states in humans. This is particularly evident in pacing myocytes and nerve terminals related to synaptic transmission. A common thread through this dissertation is on the role of three regulators proteins that are common to many cell types. These are the plasmalemmal Na⁺/Ca²⁺ exchanger (NCX), the Ca²⁺-ATPase (PMCA) …

Monitoring Activity Of Drosophila Larvae: Impedance & Video Microscopy Measures, Ann Simone Cooper, Robin L. Cooper

Biology Faculty Publications

No abstract provided.

Monitoring Heart Rate In Drosophila Larvae By Various Approaches, Sameera Dasari, Robin L. Cooper

Biology Faculty Publications

No abstract provided.

Monitoring Carbon Dioxide Production By Drosophila Larvae, Robin L. Cooper, D. Nicholas Mcletchie

Biology Faculty Publications

No abstract provided.

Drosophila Unpaired Encodes A Secreted Protein That Activates The Jak Signaling Pathway, Douglas A. Harrison, Patricia E. Mccoon, Richard Binari, Michael Gilman, Norbert Perrimon

Biology Faculty Publications

In vertebrates, many cytokines and growth factors have been identified as activators of the JAK/STAT signaling pathway. In Drosophila, JAK and STAT molecules have been isolated, but no ligands or receptors capable of activating the pathway have been described. We have characterized the unpaired (upd) gene, which displays the same distinctive embryonic mutant defects as mutations in the Drosophila JAK (hopscotch) and STAT (stat92E) genes. Upd is a secreted protein, associated with the extracellular matrix, that activates the JAK pathway. We propose that Upd is a ligand that relies on JAK signaling to stimulate transcription of pair-rule genes in a …