Olivia Sheppard | University College London (original) (raw)

Papers by Olivia Sheppard

Research paper thumbnail of Massively Parallel Sequencing Reveals the Complex Structure of an Irradiated Human Chromosome on a Mouse Background in the Tc1 Model of Down Syndrome

Research paper thumbnail of Mouse models of aneuploidy

Research paper thumbnail of Altered regulation of tau phosphorylation in a mouse model of down syndrome aging

Down syndrome (DS) results from trisomy of human chromosome 21 (Hsa21) and is associated with an ... more Down syndrome (DS) results from trisomy of human chromosome 21 (Hsa21) and is associated with an increased risk of Alzheimer's disease (AD). Here, using the unique transchromosomic Tc1 mouse model of DS we investigate the influence of trisomy of Hsa21 on the protein tau, which is hyperphosphorylated in Alzheimer's disease. We show that in old, but not young, Tc1 mice increased phosphorylation of tau occurs at a site suggested to be targeted by the Hsa21 encoded kinase, dual-specificity tyrosine-(Y)-phosphorylation regulated kinase 1A (DYRK1A). We show that DYRK1A is upregulated in young and old Tc1 mice, but that young trisomic mice may be protected from accumulating aberrantly phosphorylated tau. We observe that the key tau kinase, glycogen synthase kinase3-β (GSK-3β) is aberrantly phosphorylated at an inhibitory site in the aged Tc1 brain which may reduce total glycogen synthase kinase3-β activity. It is possible that a similar mechanism may also occur in people with DS.Copyright © 2012 Elsevier Inc. All rights reserved.

Research paper thumbnail of (‐) Epicatechin Stimulates ERK‐dependent Cyclic AMP Response Element Activity and Up‐regulates GluR2 In Cortical Neurons

Journal of …, Jan 1, 2007

Research paper thumbnail of Positive Regulation of the NADPH Oxidase NOX4 Promoter In Vascular Smooth Muscle Cells by E2F

Free Radical Biology and …, Jan 1, 2008

The generation of reactive oxygen species (ROS) by the NOX family of NADPH oxidases is known to b... more The generation of reactive oxygen species (ROS) by the NOX family of NADPH oxidases is known to be involved in the regulation of many physiological cellular functions. Unlike other members of this family, NOX4 generates ROS constitutively without the need for a stimulus. The activity of NOX4 is known to be regulated, at least in part, at the level of mRNA expression. However, nothing is known of the molecular mechanisms which underlie its transcriptional regulation. We have therefore determined the transcriptional initiation site of NOX4 in vascular smooth muscle cells (VSMC) and identified NOX4 genomic sequences necessary to effect high levels of expression of a linked luciferase reporter gene in both rat and mouse VSMCs. A potential binding site for members of the E2F family of transcription factors was identified, and electrophoretic mobility-shift assays (EMSA) and chromatin immunoprecipitation (ChIP) assays confirmed that this site binds E2F1 both in vitro and in vivo. siRNA against E2F1 decreased NOX4 promoter activity, while site-specific mutation of the core-binding site both downregulated the NOX4 promoter and abolished transregulation by E2F1. These data therefore demonstrate that E2F factor(s) are positive regulators of NOX4 transcription in VSMCs.

Research paper thumbnail of Generation of a Panel of Antibodies Against Proteins Encoded on Human Chromosome 21

Journal of Negative …, Jan 1, 2010

BackgroundDown syndrome (DS) is caused by trisomy of all or part of chromosome 21. To further und... more BackgroundDown syndrome (DS) is caused by trisomy of all or part of chromosome 21. To further understanding of DS we are working with a mouse model, the Tc1 mouse, which carries most of human chromosome 21 in addition to the normal mouse chromosome complement. This mouse is a model for human DS and recapitulates many of the features of the human syndrome such as specific heart defects, and cerebellar neuronal loss. The Tc1 mouse is mosaic for the human chromosome such that not all cells in the model carry it. Thus to help our investigations we aimed to develop a method to identify cells that carry human chromosome 21 in the Tc1 mouse. To this end, we have generated a panel of antibodies raised against proteins encoded by genes on human chromosome 21 that are known to be expressed in the adult brain of Tc1 miceResultsWe attempted to generate human specific antibodies against proteins encoded by human chromosome 21. We selected proteins that are expressed in the adult brain of Tc1 mice and contain regions of moderate/low homology with the mouse ortholog. We produced antibodies to seven human chromosome 21 encoded proteins. Of these, we successfully generated three antibodies that preferentially recognise human compared with mouse SOD1 and RRP1 proteins on western blots. However, these antibodies did not specifically label cells which carry a freely segregating copy of Hsa21 in the brains of our Tc1 mouse model of DS.ConclusionsAlthough we have successfully isolated new antibodies to SOD1 and RRP1 for use on western blots, in our hands these antibodies have not been successfully used for immunohistochemistry studies. These antibodies are freely available to other researchers. Our data high-light the technical difficulty of producing species-specific antibodies for both western blotting and immunohistochemistry.

Research paper thumbnail of Down Syndrome and the Molecular Pathogenesis Resulting From Trisomy of Human Chromosome 21

Journal of Biomedical …, Jan 1, 2010

Research paper thumbnail of Massively Parallel Sequencing Reveals the Complex Structure of an Irradiated Human Chromosome on a Mouse Background in the Tc1 Model of Down Syndrome

Research paper thumbnail of Mouse models of aneuploidy

Research paper thumbnail of Altered regulation of tau phosphorylation in a mouse model of down syndrome aging

Down syndrome (DS) results from trisomy of human chromosome 21 (Hsa21) and is associated with an ... more Down syndrome (DS) results from trisomy of human chromosome 21 (Hsa21) and is associated with an increased risk of Alzheimer's disease (AD). Here, using the unique transchromosomic Tc1 mouse model of DS we investigate the influence of trisomy of Hsa21 on the protein tau, which is hyperphosphorylated in Alzheimer's disease. We show that in old, but not young, Tc1 mice increased phosphorylation of tau occurs at a site suggested to be targeted by the Hsa21 encoded kinase, dual-specificity tyrosine-(Y)-phosphorylation regulated kinase 1A (DYRK1A). We show that DYRK1A is upregulated in young and old Tc1 mice, but that young trisomic mice may be protected from accumulating aberrantly phosphorylated tau. We observe that the key tau kinase, glycogen synthase kinase3-β (GSK-3β) is aberrantly phosphorylated at an inhibitory site in the aged Tc1 brain which may reduce total glycogen synthase kinase3-β activity. It is possible that a similar mechanism may also occur in people with DS.Copyright © 2012 Elsevier Inc. All rights reserved.

Research paper thumbnail of (‐) Epicatechin Stimulates ERK‐dependent Cyclic AMP Response Element Activity and Up‐regulates GluR2 In Cortical Neurons

Journal of …, Jan 1, 2007

Research paper thumbnail of Positive Regulation of the NADPH Oxidase NOX4 Promoter In Vascular Smooth Muscle Cells by E2F

Free Radical Biology and …, Jan 1, 2008

The generation of reactive oxygen species (ROS) by the NOX family of NADPH oxidases is known to b... more The generation of reactive oxygen species (ROS) by the NOX family of NADPH oxidases is known to be involved in the regulation of many physiological cellular functions. Unlike other members of this family, NOX4 generates ROS constitutively without the need for a stimulus. The activity of NOX4 is known to be regulated, at least in part, at the level of mRNA expression. However, nothing is known of the molecular mechanisms which underlie its transcriptional regulation. We have therefore determined the transcriptional initiation site of NOX4 in vascular smooth muscle cells (VSMC) and identified NOX4 genomic sequences necessary to effect high levels of expression of a linked luciferase reporter gene in both rat and mouse VSMCs. A potential binding site for members of the E2F family of transcription factors was identified, and electrophoretic mobility-shift assays (EMSA) and chromatin immunoprecipitation (ChIP) assays confirmed that this site binds E2F1 both in vitro and in vivo. siRNA against E2F1 decreased NOX4 promoter activity, while site-specific mutation of the core-binding site both downregulated the NOX4 promoter and abolished transregulation by E2F1. These data therefore demonstrate that E2F factor(s) are positive regulators of NOX4 transcription in VSMCs.

Research paper thumbnail of Generation of a Panel of Antibodies Against Proteins Encoded on Human Chromosome 21

Journal of Negative …, Jan 1, 2010

BackgroundDown syndrome (DS) is caused by trisomy of all or part of chromosome 21. To further und... more BackgroundDown syndrome (DS) is caused by trisomy of all or part of chromosome 21. To further understanding of DS we are working with a mouse model, the Tc1 mouse, which carries most of human chromosome 21 in addition to the normal mouse chromosome complement. This mouse is a model for human DS and recapitulates many of the features of the human syndrome such as specific heart defects, and cerebellar neuronal loss. The Tc1 mouse is mosaic for the human chromosome such that not all cells in the model carry it. Thus to help our investigations we aimed to develop a method to identify cells that carry human chromosome 21 in the Tc1 mouse. To this end, we have generated a panel of antibodies raised against proteins encoded by genes on human chromosome 21 that are known to be expressed in the adult brain of Tc1 miceResultsWe attempted to generate human specific antibodies against proteins encoded by human chromosome 21. We selected proteins that are expressed in the adult brain of Tc1 mice and contain regions of moderate/low homology with the mouse ortholog. We produced antibodies to seven human chromosome 21 encoded proteins. Of these, we successfully generated three antibodies that preferentially recognise human compared with mouse SOD1 and RRP1 proteins on western blots. However, these antibodies did not specifically label cells which carry a freely segregating copy of Hsa21 in the brains of our Tc1 mouse model of DS.ConclusionsAlthough we have successfully isolated new antibodies to SOD1 and RRP1 for use on western blots, in our hands these antibodies have not been successfully used for immunohistochemistry studies. These antibodies are freely available to other researchers. Our data high-light the technical difficulty of producing species-specific antibodies for both western blotting and immunohistochemistry.

Research paper thumbnail of Down Syndrome and the Molecular Pathogenesis Resulting From Trisomy of Human Chromosome 21

Journal of Biomedical …, Jan 1, 2010