A Feasibility Study Concerning School District Reoganization in Southern Dupage County, Illinois

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Dysregulation of the Norepinephrine Transporter Sustains Cortical Hypodopaminergia and Schizophrenia-Like Behaviors in Neuronal Rictor Null Mice

A novel animal model highlights the link between Akt dysfunction, reduced cortical dopamine function, norepinephrine transporters, and schizophrenia-like behaviors

Expression of Genes Encoding Multi-Transmembrane Proteins in Specific Primate Taste Cell Populations

BACKGROUND: Using fungiform (FG) and circumvallate (CV) taste buds isolated by laser capture microdissection and analyzed using gene arrays, we previously constructed a comprehensive database of gene expression in primates, which revealed over 2,300 taste bud-associated genes. Bioinformatics analyses identified hundreds of genes predicted to encode multi-transmembrane domain proteins with no previous association with taste function. A first step in elucidating the roles these gene products play in gustation is to identify the specific taste cell types in which they are expressed. METHODOLOGY/PRINCIPAL FINDINGS: Using double label in situ hybridization analyses, we identified seven new genes expressed in specific taste cell types, including sweet, bitter, and umami cells (TRPM5-positive), sour cells (PKD2L1-positive), as well as other taste cell populations. Transmembrane protein 44 (TMEM44), a protein with seven predicted transmembrane domains with no homology to GPCRs, is expressed in a TRPM5-negative and PKD2L1-negative population that is enriched in the bottom portion of taste buds and may represent developmentally immature taste cells. Calcium homeostasis modulator 1 (CALHM1), a component of a novel calcium channel, along with family members CALHM2 and CALHM3; multiple C2 domains; transmembrane 1 (MCTP1), a calcium-binding transmembrane protein; and anoctamin 7 (ANO7), a member of the recently identified calcium-gated chloride channel family, are all expressed in TRPM5 cells. These proteins may modulate and effect calcium signalling stemming from sweet, bitter, and umami receptor activation. Synaptic vesicle glycoprotein 2B (SV2B), a regulator of synaptic vesicle exocytosis, is expressed in PKD2L1 cells, suggesting that this taste cell population transmits tastant information to gustatory afferent nerve fibers via exocytic neurotransmitter release. CONCLUSIONS/SIGNIFICANCE: Identification of genes encoding multi-transmembrane domain proteins expressed in primate taste buds provides new insights into the processes of taste cell development, signal transduction, and information coding. Discrete taste cell populations exhibit highly specific gene expression patterns, supporting a model whereby each mature taste receptor cell is responsible for sensing, transmitting, and coding a specific taste quality

Synphilin-1 Enhances α-Synuclein Aggregation in Yeast and Contributes to Cellular Stress and Cell Death in a Sir2-Dependent Manner

© 2010 Büttner et al. This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.Background: Parkinson’s disease is characterized by the presence of cytoplasmic inclusions, known as Lewy bodies, containing both aggregated α-synuclein and its interaction partner, synphilin-1. While synphilin-1 is known to accelerate inclusion formation by α-synuclein in mammalian cells, its effect on cytotoxicity remains elusive. Methodology/Principal Findings: We expressed wild-type synphilin-1 or its R621C mutant either alone or in combination with α-synuclein in the yeast Saccharomyces cerevisiae and monitored the intracellular localization and inclusion formation of the proteins as well as the repercussions on growth, oxidative stress and cell death. We found that wild-type and mutant synphilin-1 formed inclusions and accelerated inclusion formation by α-synuclein in yeast cells, the latter being correlated to enhanced phosphorylation of serine-129. Synphilin-1 inclusions co-localized with lipid droplets and endomembranes. Consistently, we found that wild-type and mutant synphilin-1 interacts with detergent-resistant membrane domains, known as lipid rafts. The expression of synphilin-1 did not incite a marked growth defect in exponential cultures, which is likely due to the formation of aggresomes and the retrograde transport of inclusions from the daughter cells back to the mother cells. However, when the cultures approached stationary phase and during subsequent ageing of the yeast cells, both wild-type and mutant synphilin-1 reduced survival and triggered apoptotic and necrotic cell death, albeit to a different extent. Most interestingly, synphilin-1 did not trigger cytotoxicity in ageing cells lacking the sirtuin Sir2. This indicates that the expression of synphilin-1 in wild-type cells causes the deregulation of Sir2-dependent processes, such as the maintenance of the autophagic flux in response to nutrient starvation. Conclusions/Significance: Our findings demonstrate that wild-type and mutant synphilin-1 are lipid raft interacting proteins that form inclusions and accelerate inclusion formation of α-synuclein when expressed in yeast. Synphilin-1 thereby induces cytotoxicity, an effect most pronounced for the wild-type protein and mediated via Sir2-dependent processes.This work was supported by grants from IWT-Vlaanderen (SBO NEURO-TARGET), the K.U.Leuven Research Fund (K.U.Leuven BOF-IOF) and K.U.Leuven R&D to JW, a Tournesol grant from Egide (Partenariat Hubert Curien) in France in collaboration with the Flemish Ministry of Education and the Fund of Scientific Research of Flanders (FWO) in Belgium to JW, MCG and LB, a shared PhD fellowship of the EU-Marie Curie PhD Graduate School NEURAD to JW, MCG and LB, grants of the Austrian Science Fund FWF (Austria) to FM and DR (S-9304-B05), to FM and SB (LIPOTOX), and to SB (T-414-B09; Hertha-Firnberg Fellowship) and an EMBO Installation Grant, a Marie Curie IRG, and a grant of the Fundação para a Ciência e Tecnologia (PTDC/SAU-NEU/105215/2008) to TFO. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript

Transgenic <i>Xenopus laevis</i> Line for In Vivo Labeling of Nephrons within the Kidney.

Xenopus laevis embryos are an established model for studying kidney development. The nephron structure and genetic pathways that regulate nephrogenesis are conserved between Xenopus and humans, allowing for the study of human disease-causing genes. Xenopus embryos are also amenable to large-scale screening, but studies of kidney disease-related genes have been impeded because assessment of kidney development has largely been limited to examining fixed embryos. To overcome this problem, we have generated a transgenic line that labels the kidney. We characterize this cdh17:eGFP line, showing green fluorescent protein (GFP) expression in the pronephric and mesonephric kidneys and colocalization with known kidney markers. We also demonstrate the feasibility of live imaging of embryonic kidney development and the use of cdh17:eGFP as a kidney marker for secretion assays. Additionally, we develop a new methodology to isolate and identify kidney cells for primary culture. We also use morpholino knockdown of essential kidney development genes to establish that GFP expression enables observation of phenotypes, previously only described in fixed embryos. Taken together, this transgenic line will enable primary kidney cell culture and live imaging of pronephric and mesonephric kidney development. It will also provide a simple means for high-throughput screening of putative human kidney disease-causing genes

Tissue-Specific Gene Inactivation in Xenopus laevis : Knockout of lhx1 in the Kidney with CRISPR/Cas9

Author Posting. © The Author(s), 2017. This is the author's version of the work. It is posted here by permission of Genetics Society of America for personal use, not for redistribution. The definitive version was published in Genetics 208 (2018): 673-686, doi:10.1534/genetics.117.300468.Studying genes involved in organogenesis is often difficult because many of these genes are also essential for early development. The allotetraploid frog, Xenopus laevis, is commonly used to study developmental processes, but because of the presence of two homeologs for many genes, it has been difficult to use as a genetic model. Few studies have successfully used CRISPR in amphibians, and currently there is no tissue-targeted knockout strategy described in Xenopus. The goal of this study is to determine whether CRISPR/Cas9-mediated gene knockout can be targeted to the Xenopus kidney without perturbing essential early gene function. We demonstrate that targeting CRISPR gene editing to the kidney and the eye of F0 embryos is feasible. Our study shows that knockout of both homeologs of lhx1 results in the disruption of kidney development and function but does not lead to early developmental defects. Therefore, targeting of CRISPR to the kidney may not be necessary to bypass the early developmental defects reported upon disruption of Lhx1 protein expression or function by morpholinos, antisense RNA, or dominant negative constructs. We also establish a control for CRISPR in Xenopus by editing a gene (slc45a2) that when knocked out results in albinism without altering kidney development. This study establishes the feasibility of tissue-specific gene knockout in Xenopus, providing a cost effective and efficient method for assessing the roles of genes implicated in developmental abnormalities that is amenable to high-throughput gene or drug screening techniques.These studies were supported by a National Institutes of Health (NIH) KO1 grant (K01DK092320 to R.K.M.), startup funding from the Department of Pediatrics 424 Pediatric Research Center at the University of Texas McGovern Medical School (to R.K.M.), an NIH National Xenopus Resource Center grant (P40OD010997 to M.E.H.), and an NIH R01 grant (R01HD084409 to M.E.H.)