1991 — 1993 |
George, Alfred L |
K08Activity Code Description: To provide the opportunity for promising medical scientists with demonstrated aptitude to develop into independent investigators, or for faculty members to pursue research aspects of categorical areas applicable to the awarding unit, and aid in filling the academic faculty gap in these shortage areas within health profession's institutions of the country. |
Human Muscle Sodium Channels in Period Paralysis
The research proposal within this application will use a molecular genetic approach to examine the hypothesis that a subpopulation of abnormal voltage-gated sodium channels expressed in skeletal muscle is responsible for the pathogenesis of the hereditary periodic paralyses. The hereditary periodic paralyses are a heterogenous group of nondystrophic autosomal dominant muscle diseases in which episodic failure of muscle membranes to generate and propagate action potentials leads to intermittent attacks of weakness or flaccid paralysis. Experimental evidence suggests that a noninactivating sodium conductance in surface membranes, possibly arising from a subpopulation of mutant sodium channels, leads to membrane depolarization and inactivation of normal voltage-gated sodium channels. The specific aims of this proposal are to clone and sequence the predominant human muscle voltage-gated sodium channels and identify restriction fragment length polymorphisms (RFLP) for use in chromosome mapping and linkage analysis in families with periodic paralysis. This information will lead to an improved understanding of the pathogenesis of this intriguing group of genetic muscle disorders, and help characterize the structure of a protein essential to the contraction of human striated muscle.
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0.915 |
1994 — 1995 |
George, Alfred L |
K08Activity Code Description: To provide the opportunity for promising medical scientists with demonstrated aptitude to develop into independent investigators, or for faculty members to pursue research aspects of categorical areas applicable to the awarding unit, and aid in filling the academic faculty gap in these shortage areas within health profession's institutions of the country. |
Muscle Sodium Channels in Period Paralysis
The research proposal within this application will use a molecular genetic approach to examine the hypothesis that a subpopulation of abnormal voltage-gated sodium channels expressed in skeletal muscle is responsible for the pathogenesis of the hereditary periodic paralyses. The hereditary periodic paralyses are a heterogenous group of nondystrophic autosomal dominant muscle diseases in which episodic failure of muscle membranes to generate and propagate action potentials leads to intermittent attacks of weakness or flaccid paralysis. Experimental evidence suggests that a noninactivating sodium conductance in surface membranes, possibly arising from a subpopulation of mutant sodium channels, leads to membrane depolarization and inactivation of normal voltage-gated sodium channels. The specific aims of this proposal are to clone and sequence the predominant human muscle voltage-gated sodium channels and identify restriction fragment length polymorphisms (RFLP) for use in chromosome mapping and linkage analysis in families with periodic paralysis. This information will lead to an improved understanding of the pathogenesis of this intriguing group of genetic muscle disorders, and help characterize the structure of a protein essential to the contraction of human striated muscle.
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0.915 |
1994 — 2015 |
George, Alfred L |
R01Activity Code Description: To support a discrete, specified, circumscribed project to be performed by the named investigator(s) in an area representing his or her specific interest and competencies. R37Activity Code Description: To provide long-term grant support to investigators whose research competence and productivity are distinctly superior and who are highly likely to continue to perform in an outstanding manner. Investigators may not apply for a MERIT award. Program staff and/or members of the cognizant National Advisory Council/Board will identify candidates for the MERIT award during the course of review of competing research grant applications prepared and submitted in accordance with regular PHS requirements. |
Hereditary Defects in Human Sodium Channels
DESCRIPTION (provided by applicant): Voltage-gated sodium channels are heteromultimeric integral membrane proteins that are responsible for the initial phase of the action potential in most excitable cells. A variety of inherited disorders affecting skeletal muscle contraction (hyperkalemic periodic paralysis, paramyotonia congenita, K+-aggravated myotonia), cardiac excitability (congenital long QT syndrome, idiopathic ventricular fibrillation, familial conduction system disease) and certain forms of epilepsy have been associated with mutations in various human sodium channel genes. This proposal is a competing renewal of R01-NS32387 that for 8 years has funded our efforts to elucidate the molecular genetic, physiologic and pharmacologic mechanisms of human sodium "channelopathies". We have recently shifted our focus from studies of the two striated muscle sodium channel genes (SCN4A, SCN5A) to investigations of brain sodium channel genes and their role in inherited epilepsies. We propose to perform a series of carefully integrated experiments employing molecular genetic, recombinant DNA and cellular electrophysiological approaches to elucidate the molecular defects responsible for seizure disorders linked to three distinct neuronal sodium channel genes (SCN1B, SCNIA, SCN2A). In Specific Aim 1, we propose to perform molecular genetic screening in a large cohort of families segregating seizure phenotypes consistent with generalized epilepsy with febrile seizures plus (GEFS+), severe myoelonic epilepsy of infancy (SMEI) and other less well characterized disorders that may be associated with mutations in brain sodium channels. In Specific Aim 2, we plan to perform biophysical and pharmacological characterization of epilepsy-associated mutations using recombinant human neuronal sodium channels expressed heterologously in mammalian cells. Our laboratory is uniquely qualified to elucidate the molecular mechanism of SCN1A-associated epilepsy using recombinant human SCN1A, a reagent that we have recently developed. Finally in Specific Aim 3, we will elucidate the molecular mechanisms responsible for dysfunction of the human sodium channel [31 subunit in some forms of familial epilepsy. Altogether, this work is designed to establish important correlations between genotype, clinical phenotype and biophysical properties of mutant sodium channels in human epilepsies and will have important pathophysiologic and therapeutic implications for hereditary disorders of sodium channels.
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0.915 |
1997 — 2001 |
George, Alfred L |
R01Activity Code Description: To support a discrete, specified, circumscribed project to be performed by the named investigator(s) in an area representing his or her specific interest and competencies. |
Hereditary Defects in Human Chloride Channels
DESCRIPTION: The aims of this proposal are directed toward the understanding of the structure and function of the CIC family of voltage-gated chloride channels. Mutations within the gene encoding the CIC-1 channel in human or goat skeletal muscle have been shown by a number of groups, including the applicant's, to result in inherited forms of myotonia congenita. The applicant has also contributed to the analysis of how these mutations affect channel function by characterizing wild-type and mutant channels in transfected HEK293 cells using electrophysiological methods. These naturally occurring mutations have provided a framework on which to generate further mutations by site-directed methods with the aim of mapping channel domains that constitute the voltage sensor and ion pore. In addition, the applicant proposes to delineate the transmembrane topology of the CIC channel using a glycosylation tagging method that has previously been employed to generate a topological model of AMPA-type glutamate receptors.
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0.915 |
1997 — 2001 |
George, Alfred L |
P01Activity Code Description: For the support of a broadly based, multidisciplinary, often long-term research program which has a specific major objective or a basic theme. A program project generally involves the organized efforts of relatively large groups, members of which are conducting research projects designed to elucidate the various aspects or components of this objective. Each research project is usually under the leadership of an established investigator. The grant can provide support for certain basic resources used by these groups in the program, including clinical components, the sharing of which facilitates the total research effort. A program project is directed toward a range of problems having a central research focus, in contrast to the usually narrower thrust of the traditional research project. Each project supported through this mechanism should contribute or be directly related to the common theme of the total research effort. These scientifically meritorious projects should demonstrate an essential element of unity and interdependence, i.e., a system of research activities and projects directed toward a well-defined research program goal. |
Genetic Susceptibility in the Acquired Long Qt Syndrome
One of the most striking manifestations of variability in cardiac electrophysiology is the apparently unpredictable development of the drug-induced or ~acquired~ long QT syndrome (ALQT) during antiarrhythmic therapy. ALQT, which can be fatal, occurs in up to 5% of exposed patients, even when risk factors such as pre- existing QT prolongation or hypokalemia are eliminated. The Project will combine clinical and molecular approaches to test the hypothesis that a subset of patients with ALQT is genetically predisposed to developing this adverse reaction to drug therapy. In Specific Aim 1, evidence for familial aggregation of ALQT susceptibility will be sought by challenging first degree relatives of ALQT probands with a low dose infusion of the action potential prolonging agent, ibutilide. The extent of QT prolongation in this test population will be compared with that in a control group, first degree relatives of patients who have tolerated chronic antiarrhythmic therapy. Recent advances in defining loci responsible for causing the congenital long QT syndrome have identified three cardiac ion channel genes (HERG, KvLQT1, SCN5A) as logical candidates for an ALQT susceptibility gene. Other cardiac ion channel genes (Kv1.5, Kv4.3, minK) that appear to have important roles in ventricular repolarization also candidates for this search. Specific Aim 2 will test the hypothesis that structural polymorphisms or allelic variation in DNA regulatory elements that segregate with the ALQT phenotype are present in these candidate genes; these experiments will also define the 5' end of the KvLQT1 coding sequence. Since variability in repolarization may also be acquired through variability in expression of ion channel genes, the goal of Specific Aim 3 is to identify transcriptional control sequences in HERG and in KvLQT1. The outcome of these studies will be a test of the concept that genetic factors play an important role in ALQT, and improved understanding of the molecular mechanisms determining variability in cardiac repolarization in human subjects.
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0.915 |
1997 — 2001 |
George, Alfred L |
P01Activity Code Description: For the support of a broadly based, multidisciplinary, often long-term research program which has a specific major objective or a basic theme. A program project generally involves the organized efforts of relatively large groups, members of which are conducting research projects designed to elucidate the various aspects or components of this objective. Each research project is usually under the leadership of an established investigator. The grant can provide support for certain basic resources used by these groups in the program, including clinical components, the sharing of which facilitates the total research effort. A program project is directed toward a range of problems having a central research focus, in contrast to the usually narrower thrust of the traditional research project. Each project supported through this mechanism should contribute or be directly related to the common theme of the total research effort. These scientifically meritorious projects should demonstrate an essential element of unity and interdependence, i.e., a system of research activities and projects directed toward a well-defined research program goal. |
Core--Scientific
The Scientific Core will provide oligonucleotide synthesis and DNA sequencing. An oligonucleotide synthesizer is already in place, and a DNA sequencer will be purchased. All Projects in the Program will make extensive use of both procedures for applications such as the site-directed mutagenesis, cloning and genetic mutation analysis. Centralizing these capabilities in a Core facility offers not only economies of scale, but also rapid turnaround to allow the experiments to go forward in a timely fashion.
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0.915 |
1998 |
George, Alfred L |
R13Activity Code Description: To support recipient sponsored and directed international, national or regional meetings, conferences and workshops. |
Conference--Hereditary Dysfunction of Ion Channels
DESCRIPTION (Abstract of the application) Since 1989 there has been a growing number of hereditary syndromes identified that are caused by genetic mutations in genes encoding ion channels. At the present time, there are approximately 30 identified human genetic conditions that have been linked to approximately 25 different ion channel genes. In the past three years alone more than half of the currently identified ion channel diseases have been recognized. In view of the great potential in discovery of new ion channels and ion channels, there is strong interest among a variety of biomedical scientists to examine this field in-depth. Support is requested to partially underwrite the costs of a conference titled "ION CHANNELOPATHIES: HEREDITARY DYSFUNCTION OF ION CHANNELS" to be held October 28-31, 1998 immediately following the Annual Meeting of the American Society of Nephrology. The general purpose of this conference will be to assemble the top biomedical scientists in this field along with promising young investigators and trainees to examine recent advances in the discovery of ion channel syndromes. To facilitate efforts to recruit, nurture, and develop new researchers in this field, the conference will offer travel grants to young investigators and trainees. Funds are requested in this application for 25 travel grants ($850 each) to be awarded exclusively to advanced trainees and scientists in the early phase of their careers with special consideration give to women, racial/ethnic minorities, individuals with disabilities, and others who are under-represented in science. Two posters sessions will provide a forum for these individuals to present and discuss their work with leaders in the field. The planning committee for the conference will select the awardees, and the applicant organization will serve as the fiscal agent for awarding the travel grants. The conference will be publicized in scientific journals, on the INTERNET, and announcements will be mailed to members of the sponsoring organizations (American and International Societies of Nephrology) as well as individuals active in the field. Electronic publicity will be accomplished by listing the meeting on the worldwide web home pages of the two sponsoring organizations, and by establishing a conference web site providing detains of the program and application procedure. The International Society of Nephrology has agreed to publish the proceedings of the conference as a supplemental issue of Kidney International as one of its Forefronts in Nephrology series.
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0.915 |
2000 — 2002 |
George, Alfred L |
U24Activity Code Description: To support research projects contributing to improvement of the capability of resources to serve biomedical research. |
Vanderbilt Niddk Biotechnology Center
DESCRIPTION (Abstract of the application) Vanderbilt University School of Medicine has a long history of excellence in NIDDK-supported research in diabetes, renal biology and disease, and digestive disorders. We currently enjoy a scientifically diverse portfolio of NIDDK programs including a George O'Brien Kidney Disease Research Center in its 14th year of existence, and the Vanderbilt Diabetes Research and Training Center established originally in 1978, and a total of 58 NIDDK-funded investigators in 10 different departments. This proposal seeks to build on our solid foundation of research support to establish the Vanderbilt NIDDK Biotechnology Center for enabling NIDDK-funded investigators to utilize state-of-the-art gene profiling techniques. This initiative has been made feasible by the recent creation of a microarray fabrication and analysis facility, the Vanderbilt Microarray Shared Resource. We are seeking support to expand the capabilities of this facility to provide microarray technology access to NIDDK-supported investigators at our institution. We have assembled a multidisciplinary team including faculty expert in molecular biology, bioengineering, bioinformatics, and genetics to provide direct assistance to NIDDK-funded investigators wishing to utilize this state-of-the-art gene expression technology. Support of the microarray fabrication facility will be accompanied by creation of a Data Analysis and Bioinformatics core group to assist microarray users with the task of analyzing the resulting complex data. An Internal Advisory Committee has been established to provide oversight and guidance relating to appropriate use of microarray resources, quality control, resources sharing, and in development or acquisition of new technologies. We also describe plans for establishing a Pilot and Feasibility grant program to help investigators explore new uses of cDNA microarrays for gene expression profiling, for development or testing of new methodology, and for development of new resources such as specialized libraries. To enhance awareness of the utility of microarray gene expression tools and to disseminate information about the core resources supported by this Center, we will provide training and education to investigators on the optimal use of microarray technology. Finally, we will implement plans for the timely sharing of new experimental approaches, technologies, and resources to the broader research community.
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0.915 |
2000 — 2002 |
George, Alfred L |
P01Activity Code Description: For the support of a broadly based, multidisciplinary, often long-term research program which has a specific major objective or a basic theme. A program project generally involves the organized efforts of relatively large groups, members of which are conducting research projects designed to elucidate the various aspects or components of this objective. Each research project is usually under the leadership of an established investigator. The grant can provide support for certain basic resources used by these groups in the program, including clinical components, the sharing of which facilitates the total research effort. A program project is directed toward a range of problems having a central research focus, in contrast to the usually narrower thrust of the traditional research project. Each project supported through this mechanism should contribute or be directly related to the common theme of the total research effort. These scientifically meritorious projects should demonstrate an essential element of unity and interdependence, i.e., a system of research activities and projects directed toward a well-defined research program goal. |
Functional Biology of Cic Chloride Channel in C Elegans
Voltage-gated chloride channels belonging to the CIC gene family have recently been identified as important molecular components of various physiological processes including sarcolemmal excitation, cell volume regulation, organellar acidification, and renal epithelial C1-transport. In mammalian genomes, there have been 9 distinct isoforms identified through molecular cloning, and three of the genes encoding different human CIC channels are responsible for distinct inherited diseases. Despite the clear importance of certain CIC channels, most isoforms have no known physiological function in mammalian tissues. This stems largely from the difficulty in studying these channels in complex organisms. In the complete C. elegans genome, six predicted coding sequences have been identified that exhibit homology with the mammalian CIC gene family. This Project seeks to characterize the primary structure, function, tissue localization, and physiological role of CIC gene family. This Project seeks to characterize the primary structure, function, tissue localization, and physiological role of CIC chloride channels in C. elegans using an integrated approach that utilizes molecular, electrophysiological, and genetic techniques. This comprehensive analysis of the role of CIC channels in such a well characterized model organisms presents unique opportunities to gain insight into the fundamental biological roles played by CIC channels. We will begin with cDNA cloning, functional expression, and determination of tissue localization of each gene (Specific Aim 1). We will next characterize the phenotypes associated with targeted gene disruption of each CIC channel in the work (Specific Aim 2) and be prepared to exploit genetic screens to help identified important interactions of CIC channels with other genes. The latter experiments may reveal the existence of accessory subunits or other protein-protein interactions important for channel function. Lastly, in Specific Aim 3, we have outlined a strategy to determine if heteromultimeric assemblies of CIC channels occur in vivo, and to understand how this impacts on the functional diversity and physiological importance of CIC channels in a multicellular organism.
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0.915 |
2001 — 2012 |
George, Alfred L |
T32Activity Code Description: To enable institutions to make National Research Service Awards to individuals selected by them for predoctoral and postdoctoral research training in specified shortage areas. |
Training Program in Ion Channel and Transporter Biology
DESCRIPTION (Adapted From The Abstract Provided By Applicant): Ion channels and transporter proteins are ubiquitous molecules that serve a variety of important physiological functions, provide targets for many types of pharmacological agents, and are encoded by genes that can be the basis for inherited diseases affecting the nervous system and other tissues. This proposal describes a new Training Program in Ion Channel and Transporter Biology that will strive to provide multidisciplinary research training for pre-doctoral and post-doctoral basic scientists and physicianscientists. This highly focused training program will involve 18 NIH-funded mentors in 7 different academic departments at Vanderbilt University with strong records of accomplishments in the ion channel and transporter field, and with a deep commitment to training students and postdoctoral fellows. Based on the breadth of research expertise of the training faculty, trainees can be expected to gain experiences in one or more of the following disciplines: physiology, pharmacology, neuroscience, molecular biology, structural biology, and genetics. Pre-doctoral students will be recruited from a national pool of applicants who apply for graduate studies in our Interdisciplinary Graduate and Medical Scientist Training Programs. Additional pre-doctoral students may emerge from our Medical Scholars Program, an innovative research leave program in the Vanderbilt Medical School. Post-doctoral trainees will be selected from the pool of applicants that apply to preceptor laboratories as well as physicianscientist applicants from various clinical training programs at the Vanderbilt University Medical Center. In addition to intensive research experiences, both pre-doctoral and post-doctoral trainees will have rigorous didactic course requirements, and have formal mentoring and career guidance. The goal of the training program is to develop basic scientists and physicianscientists with strong commitments to academic biomedical research in ion channel and transporter biology especially in areas with direct relevance to human health and disease.
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0.915 |
2001 — 2011 |
George, Alfred L |
R01Activity Code Description: To support a discrete, specified, circumscribed project to be performed by the named investigator(s) in an area representing his or her specific interest and competencies. |
Genetic Modifiers of Congenital Long Qt Syndrome
DESCRIPTION (provided by applicant): Ventricular arrhythmias remain the single most important cause of sudden cardiac death (SCD) among adults living in industrialized nations. Genetic factors have substantial effects in determining population-based risk for SCD and may also account for interindividual variability in susceptibility. Great progress has been made in identifying genes underlying various Mendelian disorders associated with inherited arrhythmia susceptibility. The most well studied familial arrhythmia syndrome is the congenital long QT syndrome (LQTS) caused by mutations in genes encoding subunits of myocardial ion channels. Not all mutation carriers have equal risk for experiencing the clinical manifestations of the disease (ie: syncope, sudden death). This observation has raised the possibility that genetic factors other than the primary disease-associated mutation can modify the risk of LQTS manifestations. This proposal outlines an international collaboration designed to investigate the role of candidate modifier genes in determining disease expression in a unique founder population of 17 LQTS families in South Africa. We hypothesize the existence of two types of modifier genes: genes which affect the magnitude of the underlying myocardial arrhythmia substrate, and genes which affect the propensity for triggering events acting through the autonomic nervous system. The primary goals of the study include further ascertainment and extension of the identified pedigrees and to assess a variety of surrogate clinical markers of autonomic function in mutation carriers (Specific Aim 1), to examine multiple candidate modifier genes for their association with symptomatic and asymptomatic disease (Specific Aim 2), and to test the hypothesis that variable transcriptional control of the primary disease gene (KCNQ1) impacts on arrhythmia susceptibility (Specific Aim 3). Identification of LQTS modifiers will enhance our understanding of the pathophysiology of LQTS, provide new and valuable information for counseling patients with this disorder, and will contribute to our understanding of more common arrhythmia syndromes associated with highly prevalent cardiac diseases (e.g. ischemic heart disease and congestive heart failure) that are burdened by a high incidence of SCD.
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0.915 |
2002 — 2005 |
George, Alfred L |
P01Activity Code Description: For the support of a broadly based, multidisciplinary, often long-term research program which has a specific major objective or a basic theme. A program project generally involves the organized efforts of relatively large groups, members of which are conducting research projects designed to elucidate the various aspects or components of this objective. Each research project is usually under the leadership of an established investigator. The grant can provide support for certain basic resources used by these groups in the program, including clinical components, the sharing of which facilitates the total research effort. A program project is directed toward a range of problems having a central research focus, in contrast to the usually narrower thrust of the traditional research project. Each project supported through this mechanism should contribute or be directly related to the common theme of the total research effort. These scientifically meritorious projects should demonstrate an essential element of unity and interdependence, i.e., a system of research activities and projects directed toward a well-defined research program goal. R21Activity Code Description: To encourage the development of new research activities in categorical program areas. (Support generally is restricted in level of support and in time.) |
Transcriptional Remodeling in Cardiac Arrhythmias
DESCRIPTION (provided by applicant): Electrical remodeling describes the electrophysiological changes occurring in chronic cardiac diseases associated with an increased susceptibility to arrhythmias. Changes reflecting remodeling in readily measured electrophysiologic characteristics (action potential duration, individual ionic currents) have been observed in large animal models with arrhythmias closely resembling those in humans. In the dog model of chronic complete AV block (CAVB), biventricular hypertrophy occurs along with substantial increases in action potential duration that render the animals prone to developing Torsades de Pointes during exposure to QT-prolonging antiarrhythmic agents. Similarly in chronic atrial fibrillation (both human and various pacing-induced animal models), atrial electrical remodeling produces a cellular substrate that perpetuates the arrhythmia (?atrial fibrillation begets atrial fibrillation?). Evidence points to long term changes in gene expression as an important element in the genesis of these arrhythmia prone states. However, our knowledge of the identity and temporal sequence of changes in gene expression underlying these two conditions is rudimentary. In this Project, we will use microarray technology to survey global patterns of transcriptional remodeling in dog hearts that occur following chronic AV block and with pacing-induced atrial fibrillation. This work requires that we develop a gene array from a panel of canine expressed sequence tags that we have started to collect. Development of a canine microarray will enable us to examine changes in gene expression in hypertrophied ventricular myocardium of CAVB dogs and assess differences between subgroups of animals that exhibit susceptibility or resistance to drug-induced Torsade de Pointes. We will also be able to characterize the transcriptional remodeling in atrial myocardium associated with induction of atrial fibrillation in dogs subjected to rapid atrial pacing. Robust statistical analyses of microarray data will be used to direct appropriate validation experiments using separate methods. These studies will provide new insights into the pathogenesis of arrhythmia susceptibility and contribute to identifying potential new targets for therapeutic interventions.
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0.915 |
2004 — 2008 |
George, Alfred L |
P30Activity Code Description: To support shared resources and facilities for categorical research by a number of investigators from different disciplines who provide a multidisciplinary approach to a joint research effort or from the same discipline who focus on a common research problem. The core grant is integrated with the center's component projects or program projects, though funded independently from them. This support, by providing more accessible resources, is expected to assure a greater productivity than from the separate projects and program projects. |
Dna Sequencing Shared Resources
The DNA Sequencing Shared Resource offers centralized, quality-controlled and automated DNA sequencing for research using molecular biological methods, recombinant DNA techniques, molecular genetics, and human genetics. Two types of DNA sequencing services are provided. In the first, Standard Sequencing Reaction and Electrophoresis, investigators provide purified DNA temples (plasmid, PCR product, BAC) and sequencing primer (or request use of a standard sequencing primer maintained by the resource) and the sequencing laboratory performs a complete sequencing reaction and electrophoresis. The investigator is then provided electronic data files containing individual sequencing results distributed by e-mail or file transfer. They are also provided with a color electropherogram of each reaction generated. Bulk DNA Sequencing Services are available to investigators who need large-scale sequencing with a limited number of sequencing primers. Dr. Alfred L. George, Jr. is the Scientific Director of the DNA Sequencing Shared Resource. He oversees all activities of the Resource including quality control, user prioritization, personnel management, and new technology. He has a capable staff with many years of experience available to offer help to VICC investigators.
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0.915 |
2005 — 2008 |
George, Alfred L |
R01Activity Code Description: To support a discrete, specified, circumscribed project to be performed by the named investigator(s) in an area representing his or her specific interest and competencies. |
Molecular Physiology of Kcne Potassium Channel Subunits
DESCRIPTION (provided by applicant): Voltage-gated potassium (Kv) channels are modulated by at least three distinct classes of accessory proteins including the KCNE family of single transmembrane proteins. In the human genome, KCNE proteins are encoded by five genes designated KCNE1 through KCNE5. The physiological importance of three KCNE genes has been demonstrated by associations with inherited disorders of cardiac and skeletal muscle excitability. Mutation of KCNE 1 impairs generation of the slowly activating cardiac delayed rectifier current (IKS), a critical potassium current contributing to myocardial repolarization that can be reconstituted in vitro by heterologous co-expression of recombinant KCNQ1 potassium channels with KCNE1. Other KCNE proteins also exert diverse functional effects on heterologously expressed KCNQ1. Co-expression of KCNQ1 with KCNE3, KCNE4 or KCNE5 is particularly interesting with functional effects ranging from constitutive activation (KCNE3) to suppression (KCNE4, KCNE5). We have recently demonstrated that all KCNE genes are expressed in human cardiac myocytes to varying degrees. We have further demonstrated that KCNE4 and KCNE5 exert potent inhibitory influences on heterologously expressed IKappaS and KCNE4 appears dominant over KCNE1 in this context. In addition to potential interactions with KCNQ1, KCNE proteins affect the functional properties of other cloned potassium channels. The physiological relevance of these in vitro observations remains uncertain. The observations that multiple KCNE proteins can modulate KCNQ1 channels to produce diverse biophysical phenotypes raises several intriguing questions with important implications for understanding the regulation of IKS as well as other potassium currents in normal and diseased heart. Can different KCNE subtypes compete functionally with KCNE1 for modulation of KCNQ1? Can KCNQ1 associate with more than one KCNE subtype in the same cell and at the same time? Do KCNE4 and KCNE5 participate in the pathophysiology of heart failure and other arrhythmia-prone conditions? In this proposal, we outline several experimental approaches for defining the physiological importance of KCNE subunits with a particular focus on KCNE4 and KCNE5. In Specific Aim 1, we will use a heterologous expression system to assess the range of functional and biochemical interactions possible between KCNQ1 and different KCNE subunits with a focus on KCNE4 and KCNE5. In Specific Aim 2, we will test the hypothesis that KCNE4 and KCNE5 are physiologically significant regulators of potassium currents in native cardiac myocytes by using two independent gene silencing approaches. Finally in Specific Aim 3, we will examine expression patterns of KCNQ1 and KCNE genes in diseased human heart and in an experimental canine model of ventricular arrhythmia susceptibility. Results from these studies will have implications for advancing our knowledge of the complex interplay between pore-forming and accessory subunits that comprise human potassium channels expressed in the heart and in other tissues.
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0.915 |
2007 — 2015 |
George, Alfred L |
R01Activity Code Description: To support a discrete, specified, circumscribed project to be performed by the named investigator(s) in an area representing his or her specific interest and competencies. |
Neonatal Long Qt Syndrome and Sudden Infant Death
[unreadable] DESCRIPTION (provided by applicant): Sudden death during the first year of life is a leading cause of infant mortality in developed countries. In neonates and infants, sudden unexplained death is classified as the sudden infant death syndrome (SIDS) when rigorous efforts to identify the cause of death including a forensic examination are unrevealing. Cardiac mechanisms including life-threatening arrhythmias are suspected to cause an undefined proportion of SIDS and recent evidence indicates that mutations in genes responsible for the congenital long QT syndrome (LQTS) are found in a significant number of cases. Molecular evidence for a link between SIDS and neonatal LOTS supports earlier observations that there is an increased risk of SIDS in infants with a QTc > 440 msec based on ECG measurements in 34,000 neonates. Additional anecdotal evidence indicates that LQTS may also present as intrauterine fetal death (IUFD). These observations emphasize the need to more fully understand the prevalence and consequences of arrhythmia-promoting genetic factors in the setting of fetal, neonatal and infant mortality. The goal of this research proposal is to advance our understanding of the genetic risks influencing susceptibility to sudden death before age 1 year. In Specific Aim 1, we will test the hypothesis that mutations in arrhythmia susceptibility genes occur in a measurable subset of SIDS victims and unexplained intrauterine fetal death in late gestation. Separately, we will determine the molecular basis for prolonged QTc interval incidentally discovered in neonates through a massive, ongoing prospective ECG screening trial conducted in Italy. The complete coding regions and splice site sequences of KCNQ1, KCNH2, SCN5A, KCNE1, KCNE2 and coding exons of other candidate genes will be surveyed for variants in four populations, two large SIDS cohorts, a series of fetal deaths, and Italian neonatal LQTS cases identified by prospective ECG screening. In Specific Aim 2, all mutations and rare variants in arrhythmia susceptibility genes will be analyzed for their functional and/or biochemical consequences. We hypothesize that a significant proportion of rare variants in these cases will cause dysfunction of the involved ion channel consistent with impaired myocardial repolarization. Finally, in Specific Aim 3, we will test the hypothesis that unequal expression of SCN5A alleles occurs in SIDS victims and is another potential genetic mechanism (allelic imbalance) that could contribute to the pathological impact of mutations and rare variants. Results from this study will have implications for diagnosis, treatment and prevention of LQTS, SIDS and premature fetal loss. Relevance to Public Health - Sudden unexplained death is a societal burden and vexing clinical problem affecting all age groups. SIDS remains a major reason for infant mortality in the Western world. Identifying factors, including genetic, that contribute to sudden unexpected death in infants has great importance for diagnosis and treatment of preventable causes of SIDS. [unreadable] [unreadable] [unreadable] [unreadable]
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0.915 |
2011 |
George, Alfred L. |
S10Activity Code Description: To make available to institutions with a high concentration of NIH extramural research awards, research instruments which will be used on a shared basis. |
Illumina Genome Analyzer Ii
DESCRIPTION (provided by applicant): The objective of this proposal is to substantially enhance the sequencing capabilities of the established and highly productive Genome Technology Core (GTC) at Vanderbilt University Medical Center through the acquisition of an Illumina Genome Analyzer IIx next-generation DNA sequencer. The GTC was founded in 2008 as an institutional core facility managed by the Office of Personalized Medicine and tasked with supporting the genomic needs of Vanderbilt investigators and their collaborators. A single Illumina Genome Analyzer instrument, necessary staff and secondary equipment were purchased or supported with institutional funds. Since its founding, the GTC has swiftly implemented a number of technologies and applications to establish a reliable and robust high-performance sequencing service. The rapid success of this effort has resulted in the near full utilization of our single instrument leading to long wait times for new projects. This application has been developed to address two vital issues identified by the staff of the GTC. First, the major users listed in this application represent a group of heavy users of the proposed instrument (more than 75% projected utilization for 3 years) that are not currently being supported, underscoring the need for additional sequencing capacity. Second, many investigators are utilizing paired end and longer read length sequencing runs to achieve higher sequencing coverage for each sample. These longer reads and paired end approaches require substantially longer instrument run times, putting further pressure on the existing instrumentation. Together, these issues provide a strong justification for this application to support an additional Illumina instrument in the GTC. The GTC has been extremely successful in supporting a diverse group of investigators that have made use of the existing instrumentation and in implementing and optimizing technical protocols and bioinformatic tools on the Illumina platform. This success will be further enhanced by the additional instrumentation to allow the GTC to efficiently support a larger group of NIH funded investigators. The continued success of the GTC will be assured by the strong institutional commitments, and extremely proficient bioinformatic capabilities.
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0.915 |