Regulation of the Human Neuronal Nitric Oxide Synthase Gene Via Alternate Promoters

Regulation of the Human Neuronal Nitric Oxide Synthase Gene Via Alternate Promoters PDF

Author: Gregory Thomas Hartt

Publisher:

Published: 2003

Total Pages:

ISBN-13:

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Abstract: Nitric oxide is a gaseous, free radical molecule that functions in the nervous system as an atypical neurotransmitter, second messenger, vasodilator, and potent neurotoxin, depending on cell type and level of expression. Physiologically, nitric oxide is thought to mediate long-term potentiation, the cellular correlate of learning and memory formation, as well as synaptic plasticity and remodeling. Under pathologic conditions such as cerebral ischemia, NOS1 overproduces nitric oxide, which is a key mediator of excitotoxic cell death and neurodegeneration. Our laboratory discovered that multiple, individually functioning promoters (5'1 and 5'2) regulate human NOS1 gene expression. The present work describes the discovery and cloning of a novel human NOS1 promoter, 5'3, and the demonstration of its unique, developmentally regulated expression pattern in the central nervous system. Promoter-specific alternative splicing contributes to NOS1 mRNA diversity in both the 5' untranslated and coding regions of the gene. We generated several lines of transgenic mice that express reporter genes under the control of separate human NOS1 promoter complexes (PR(5'1+5'2) and PR(5'3+5'4)), and studied how these alternate promoters contribute to NOS1 expression in various physiologic and pathophysiologic states in vivo. To this end, transgene expression is documented throughout normal development, which demonstrates overlapping but distinct patterns of promoter use that contribute to the overall recapitulation of endogenous NOS1 gene expression. In a murine model of striatal neurotoxicity, systemic methamphetamine administration results in promoter-specific transcriptional activation in the transgenic mice. NOS1 enzymatic activity is stimulated in motor neuron cell bodies after peripheral transection of the facial nerve; however, the transgenic NOS1 promoter complexes are insufficient to recapitulate NOS1 activation in this model. These findings help clarify how multiple promoters and mRN A diversity contribute to the complex regulation of the human NOS1 gene, and further our general understanding of transcriptional regulation of gene expression via alternate promoters.

The Human Neuronal Nitric Oxide Synthase Gene [microform] : RNA Biology Guiding Expressional Regulation

The Human Neuronal Nitric Oxide Synthase Gene [microform] : RNA Biology Guiding Expressional Regulation PDF

Author: Derek Colin Newton

Publisher: National Library of Canada = Bibliothèque nationale du Canada

Published: 2004

Total Pages: 644

ISBN-13: 9780612917705

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Protein synthesis is the ultimate goal of gene expression and a key control point for regulation. In particular, it enables the cell to rapidly manipulate protein production without new mRNA synthesis, processing or export. The human neuronal nitric oxide synthase (nNOS) gene is an excellent model to study the regulated translation of a mRNA to protein. Numerous and distinct nNOS mRNAs are generated from a single gene locus as a result of alternate promoter usage and alternate splicing patterns. A comprehensive analysis of the structure of neuronal NOS (nNOS) mRNA species revealed NOS1 to be one of the most structurally diverse human genes described to date in terms of promoter usage. Unique exon 1 variants are utilized for transcript initiation in diverse tissues and this dependence on unique genomic regions to control transcription initiation in a cell-specific fashion burdens the transcripts with complex 5'-mRNA leader sequences. These highly structured nNOS mRNA 5'-untranslated regions (5'-UTRs) were demonstrated to have profound effects on translation in both in vitro assays and cells and to contain cis-RNA elements that modulate translational efficiency in response to changes in cellular phenotype. In addition to promoter diversity, the human nNOS gene contains an alternately spliced (AS) exon present within the 5' -UTR. Alternative splicing events that do not affect the open reading frame are distinctly uncommon in mammals; therefore we assessed its functional relevance. Transient and stable transfection methods revealed that this alternatively spliced exon acts as a potent translational repressor. Hence a unique splicing event within a 5'-UTR is demonstrated to introduce a translational control element. Importantly, this represented a newer model for the translational control in mammalian cells. Mutational analysis identified a putative stem-loop structure within this exon that acts as the functional cis-element. Subsequent RNA-EMSA analyses revealed that a specific cytoplasmic RNA-binding complex interacts with this motif. An affinity purification scheme was developed and the cytoskeletal proteins actin and alpha-actinin were identified as components of the ribonucleoprotein complex associated with this exon. This intriguing finding indicates that this exon acts as a subcellular localization signal for select nNOS mRNAs.

Index Medicus

Index Medicus PDF

Author:

Publisher:

Published: 2004

Total Pages: 1940

ISBN-13:

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Vols. for 1963- include as pt. 2 of the Jan. issue: Medical subject headings.

Tag-based Next Generation Sequencing

Tag-based Next Generation Sequencing PDF

Author: Matthias Harbers

Publisher: John Wiley & Sons

Published: 2012-02-13

Total Pages: 609

ISBN-13: 352732819X

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Tag-based approaches were originally designed to increase the throughput of capillary sequencing, where concatemers of short sequences were first used in expression profiling. New Next Generation Sequencing methods largely extended the use of tag-based approaches as the tag lengths perfectly match with the short read length of highly parallel sequencing reactions. Tag-based approaches will maintain their important role in life and biomedical science, because longer read lengths are often not required to obtain meaningful data for many applications. Whereas genome re-sequencing and de novo sequencing will benefit from ever more powerful sequencing methods, analytical applications can be performed by tag-based approaches, where the focus shifts from 'sequencing power' to better means of data analysis and visualization for common users. Today Next Generation Sequence data require powerful bioinformatics expertise that has to be converted into easy-to-use data analysis tools. The book's intention is to give an overview on recently developed tag-based approaches along with means of their data analysis together with introductions to Next-Generation Sequencing Methods, protocols and user guides to be an entry for scientists to tag-based approaches for Next Generation Sequencing.

Nitric Oxide

Nitric Oxide PDF

Author:

Publisher: Academic Press

Published: 2014-08-12

Total Pages: 483

ISBN-13: 0128004398

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First published in 1943, Vitamins and Hormones is the longest-running serial published by Academic Press. The Series provides up-to-date information on vitamin and hormone research spanning data from molecular biology to the clinic. A volume can focus on a single molecule or on a disease that is related to vitamins or hormones. A hormone is interpreted broadly so that related substances, such as transmitters, cytokines, growth factors and others can be reviewed. This volume focuses on nitric oxide. Expertise of the contributors Coverage of a vast array of subjects In depth current information at the molecular to the clinical levels Three-dimensional structures in color Elaborate signaling pathways

Molecular Mechanisms in Spermatogenesis

Molecular Mechanisms in Spermatogenesis PDF

Author: C.Y. Cheng

Publisher: Springer Science & Business Media

Published: 2009-10-24

Total Pages: 289

ISBN-13: 0387095977

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In the past thirty years, significant advances have been made in the field of reproductive biology in unlocking the molecular and biochemical events that regulate spermatogenesis in the mammalian testis. It was possible because of the unprecedented breakthroughs in molecular biology, cell biology, immunology, and biochemistry. In this book entitled, Molecular Mechanisms in Spermatogenesis, a collection of chapters has been included written by colleagues on the latest development in the field using genomic and proteomic approaches to study spermatogenesis, as well as different mechanisms and/or molecules including environmental toxicants and transcription factors that regulate and/or affect spermatogenesis. The book begins with a chapter that provides the basic concept of cellular regulation of spermatogenesis. A few chapters are also dedicated to some of the latest findings on the Sertoli cell cytoskeleton and other molecules (e.g., proteases, adhesion proteins) that regulate spermatogenesis. These chapters contain thought-provoking discussions and concepts which shall be welcomed by investigators in the field. It is obvious that many of these concepts will be updated and some may be amended in the years to come. However, they will serve as a guide and the basis for investigation by scientists in the field.

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