DOCSLIB.ORG

Molecular Analysis of Edema Factor, a Bacillus Anthracis Adenylyl Cyclase

Total Page：16

File Type：pdf, Size：1020Kb

Download full-text PDF Read full-text
Molecular Analysis of Edema Factor, a Bacillus Anthracis Adenylyl Cyclase MOLECULAR ANALYSIS OF MAMMALIAN ADENYLYL CYCLASES AND EDEMA FACTOR, A BACTERIAL ADENYLYL CYCLASE TOXIN BY Srividya Suryanarayana M.Sc, Osmania University, India, 2002 Submitted to the Graduate Degree program in Biochemistry and Biophysics and the Graduate Faculty of the University of Kansas in partial fulfillment of the requirements for the degree of Doctorate of Philosophy Dissertation Committee: ______________________________ Mark Richter, PhD (Chairperson) ______________________________ Roland Seifert, M.D., PhD ______________________________ William Picking, PhD ______________________________ Krzysztof Kuczera, PhD ______________________________ Jackob Moskovitz, PhD Date Defended: 15 th December 2008 The Dissertation Committee for Srividya Suryanarayana certifies that this is the approved version of the following dissertation: MOLECULAR ANALYSIS OF MAMMALIAN ADENYLYL CYCLASES AND EDEMA FACTOR, A BACTERIAL ADENYLYL CYCLASE TOXIN Dissertation Committee: ______________________________ Mark Richter, PhD (Chairperson) ______________________________ Roland Seifert, M.D., PhD ______________________________ William Picking, PhD ______________________________ Krzysztof Kuczera, PhD ______________________________ Jackob Moskovitz, PhD Date Approved: 15 th December 2008 Abstract Adenylyl cyclases (ACs) catalyze the conversion of ATP to cAMP, an important second messenger central to many signaling pathways. Nine different isoforms of mammalian ACs (mACs) are present, each with distinct localization, physiological function and regulatory mechanisms by activators and inhibitors. In addition to mACs, bacterial AC toxins such as edema factor (EF) from Bacillus anthracis and CyaA from Bordetella pertussis have also been identified. Following infection, the AC toxins cause a dramatic increase in cAMP levels, thereby disrupting several intracellular signaling pathways. This thesis is broadly divided into two parts and is aimed at validating the active-site nucleotide analogs of mACs and EF. The first part of the thesis focuses on understanding mAC regulation and the mechanism of interaction of mACs with fluorescent 2’, 3’-O-(2, 4, 6-Trinitrophenyl) (TNP)- nucleotides. Using purified catalytic subunits of mAC (C1/C2) as model for mACs, we have observed the binding of TNP-nucleotides to C1/C2 and the resulting conformational changes monitored by fluorescence spectroscopy. The enzymatic assays have shown that TNP-nucleotides potently inhibit C1/C2 as well as holo-AC isoforms. An isoform-selective inhibition of AC1, AC2 and AC5 by TNP-nucleotides has been reported for the first time indicating that TNP-nucleotides can serve as models for rational design of potent isoform- specific AC inhibitors. Furthermore, biophysical and biochemical analysis of 1 the effects of TNP- and 2’ (3’)-O-(N-methylanthraniloyl) (MANT) -nucleotides on the individual subunits C1 and C2 show that C1 and C2 can exist as homodimers. This homodimerization may play an important physiological role in cAMP signaling. The second part of the thesis addresses the structure-activity relationship in regulating the catalytic activity of EF and its interaction with calmodulin (CaM) using MANT derivatives of ATP and GTP as probes. Our enzymatic assays have shown that MANT-nucleotides are highly potent at inhibiting EF. MANT-nucleotides are also favorable for FRET studies indicating that our robust fluorescence assays can be used for High-Throughput Screening (HTS) of EF inhibitors. Additionally, EF-CaM interaction was probed by MANT-nucleotides. We have observed that binding and activation of EF by CaM are two independent processes in the presence and absence of calcium. Furthermore, our fluorescence assays to monitor binding of oxidized CaM to EF also indicate that methionine residues in CaM play an important role in binding to EF. 2 Acknowledgements First and foremost, I would like to thank God for blessing me with this opportunity to pursue my Ph.D in the United States of America. I thank Him for supporting me through the best and toughest years of my life. Next, I am extremely thankful to my advisors Dr. Roland Seifert and Dr.Mark Richter for their invaluable guidance, support and for sharing their expertise and research insight. Thanks for helping me make a smooth transition in the U.S and giving me a sense of direction during my doctoral studies. Thanks for believing in me during the toughest times and boosting my morale to do better every day. Thanks for helping me develop my presentation, writing and research skills and providing me with financial support to complete my PhD studies. I am deeply grateful to both of them and I wouldn’t have been able to fulfill this dream without their constant encouragement and support. I would also like to acknowledge Dr.William Picking, Dr. Krzysztof Kuczera and Dr. Jackob Moskovitz for serving on my defense committee and providing helpful suggestions. I am greatly indebted to Dr.Jackob Moskovitz for allowing me to continue my PhD research in his lab. I am also grateful to my dear lab mate Dr.Cibele Pinto (University of Kansas Medical Center) for the enthusiasm, inspiration help and a wonderful friendship, which was always there when I needed it and to Dr.Andreas Gille for initiating the projects I worked on and for all the 3 support provided. I also wish to thank Derek Oien (graduate student) for all his help in the lab. Many thanks are also due for all the collaborators at various stages of my PhD studies. I wish to thank Dr.Stephen Sprang and Dr.T.C.Mou (University of Texas Southwestern Medical Center) for providing us generously with C1/C2 and Gsa proteins. I also wish to acknowledge Dr.Wei-Jen Tang (Ben- May institute of Cancer Research, University of Chicago) for kindly providing us with EF and EF3 proteins. I would also like to thank Dr. Gerald Lushington for providing me with all the help on molecular modeling. Grant support for my research projects were provided by the American Heart Association. I don’t have enough words to describe how grateful I am to my sister Usha. Without her, my dream to become a researcher would always be unfulfilled. She has been the guiding angel in my life, helped me realize what I was capable of, encouraged me, helped me achieve my goals and to stay focused, in short, I owe her all my success. I did specially like to thank my parents (Mr. Suryanarayanan and Mrs. Janaki Suryanarayanan) for their tremendous sacrifices so that I could achieve my dream. I thank them for teaching me values to be a good person, for always being supportive of my decisions, for letting me come to the United States, thousands of miles away from them, to pursue my goal. 4 I thank all my family, my sisters (Uma, Bhagyam), my brothers-in-law (Natarajan, Ravi, Kushal and Kaushik), my nieces (Priya and Hemshikha), my in-laws (Mr.Viswanathan and Mrs. Shobha Viswanathan)for being very supportive and proud of my achievements. In particular, I would specially like to acknowledge my father-in-law Mr.Viswanathan for his constant encouragement that will always help me remain grounded and work even harder to achieve my goals. Lastly, and most importantly, I thank Karthik, my husband for being my pillar of strength without whom this journey would have been incomplete. I am extremely grateful to him for always being there for me, for believing in me and my capabilities, for all his unconditional support, inspiration, love and for his loads of patience in handling difficult situations. Many thanks are due to Ms. Geetha Vani Chittoor, Dr.Vamsee Mohan Yaganti and many others for providing emotional support and for making my stay in Kansas a very memorable one. 5 Table of Contents Abstract……………………………………………………………………………..i Acknowledgements……………………………………………………………...iii Table of contents…………………………………………………………………vi List of Tables……………………………………………………………………….x List of Figures…………………………………………………………..…………xi Chapter 1: Introduction ………………………………………………………………..1 1.1 Mammalian Adenylyl cyclases (mACs)……………………………...............2 1.1.1. AC isoforms – Tissue distribution, physiological role and potential therapeutic targets…………………………………………………………………….…………7 1.1.2. AC isoforms – regulation…………………………………………......................12 1.1.3. Catalytic mechanism of AC………………………………………......................17 1.1.4. C isoforms –Challenges, limitations and future directions of AC research…………………………………………………………….…………..….2 1.2 Bacterial Adenylyl cyclase toxin Edema Factor (EF) from Bacillus anthracis………………………………………………………………………….. 27 1.2.1. Anthrax – Infection, diagnosis,treatment and prevention……….…………….29 1.2.2. Anthrax – Biological warfare and incidence in developing countries………...33 1.2.3. Anthrax- Tripartite toxin and mechanism of toxin translocation into host cells ………………………………………………………………………..………….….35 1.2.4. Anthrax toxin components and their function…………………………………..40 1.2.5. Edema Factor – structure and intracellular activation…………………………42 1.2.6. Catalytic mechanism of EF………………………………………………………47 1.3 Challenges, limitations and future directions of anthrax research with special focus on EF…………………………………………………………………….…..51 1.4 Research presented in this thesis…………………………………………….53 6 1.5 References………………………………………………………………………...55 Chapter 2: Molecular analysis of the interaction of 2’, 3’-O-(2, 4, 6- trinitrophenyl) TNP-nucleotides with purified catalytic C1/C2 subunits of mammalian adenylyl cyclase and holo adenylyl cyclase isoforms 2.1 Introduction……………………………………………………………………………71 2.2 Specific aims and hypothesis……………………………………………………...77 2.3 Materials and methods………………………………………………………………79
Load more

Recommended publications
  • Gene and Pathway-Based Second-Wave Analysis of Genome-Wide Association Studies
    European Journal of Human Genetics (2010) 18, 111–117 & 2010 Macmillan Publishers Limited All rights reserved 1018-4813/10 $32.00 www.nature.com/ejhg ARTICLE Gene and pathway-based second-wave analysis of genome-wide association studies Gang Peng1, Li Luo2, Hoicheong Siu1, Yun Zhu1, Pengfei Hu1, Shengjun Hong1, Jinying Zhao3, Xiaodong Zhou4, John D Reveille4, Li Jin1, Christopher I Amos5 and Momiao Xiong*,2 Despite the great success of genome-wide association studies (GWAS) in identification of the common genetic variants associated with complex diseases, the current GWAS have focused on single-SNP analysis. However, single-SNP analysis often identifies only a few of the most significant SNPs that account for a small proportion of the genetic variants and offers only a limited understanding of complex diseases. To overcome these limitations, we propose gene and pathway-based association analysis as a new paradigm for GWAS. As a proof of concept, we performed a comprehensive gene and pathway-based association analysis of 13 published GWAS. Our results showed that the proposed new paradigm for GWAS not only identified the genes that include significant SNPs found by single-SNP analysis, but also detected new genes in which each single SNP conferred a small disease risk; however, their joint actions were implicated in the development of diseases. The results also showed that the new paradigm for GWAS was able to identify biologically meaningful pathways associated with the diseases, which were confirmed by a gene-set-rich analysis using gene expression
    [Show full text]
  • Microrna Let-7I-5P Mediates the Relationship Between Muscle Fat
    www.nature.com/scientificreports OPEN microRNA let‑7i‑5p mediates the relationship between muscle fat infltration and neck pain disability following motor vehicle collision: a preliminary study James M. Elliott1,2, Cathleen A. Rueckeis3, Yue Pan3,4, Todd B. Parrish2,5, David M. Walton6 & Sarah D. Linnstaedt3,7* Persistent neck‑pain disability (PNPD) is common following traumatic stress exposures such as motor vehicle collision (MVC). Substantial literature indicates that fat infltration into neck muscle (MFI) is associated with post‑MVC PNPD. However, little is known about the molecular mediators underlying this association. In the current study, we assessed whether microRNA expression signatures predict PNPD and whether microRNA mediate the relationship between neck MFI and PNPD. A nested cohort of 43 individuals from a longitudinal study of MVC survivors, who provided blood (PAXgene RNA) and underwent magnetic resonance imaging (MRI), were included in the current study. Peritraumatic microRNA expression levels were quantifed via small RNA sequencing, neck MFI via MRI, and PNPD via the Neck Disability Index two‑weeks, three‑months, and twelve‑months following MVC. Repeated measures regression models were used to assess the relationship between microRNA and PNPD and to perform mediation analyses. Seventeen microRNA predicted PNPD following MVC. One microRNA, let‑7i‑5p, mediated the relationship between neck MFI and PNPD. Peritraumatic blood‑ based microRNA expression levels predict PNPD following MVC and let‑7i‑5p might contribute to the underlying efects of neck MFI on persistent disability. In conclusion, additional studies are needed to validate this fnding. While most individuals recover following traumatic stress exposures such as motor vehicle collision (MVC), a substantial subset develop adverse posttraumatic neuropsychiatric sequelae such as persistent neck pain and persistent neck-pain related disability (PNPD)1–4, which is typically measured by a self-report region-specifc disability scale (such as the Neck Disability Index (NDI)5).
    [Show full text]
  • Therapeutic Opportunities in Colon Cancer Focus on Phosphodiesterase Inhibitors
    Life Sciences 230 (2019) 150–161 Contents lists available at ScienceDirect Life Sciences journal homepage: www.elsevier.com/locate/lifescie Review article Therapeutic opportunities in colon cancer: Focus on phosphodiesterase inhibitors T ⁎ Ankita Mehta, Bhoomika M. Patel Institute of Pharmacy, Nirma University, Ahmedabad, India ARTICLE INFO ABSTRACT Keywords: Despite novel technologies, colon cancer remains undiagnosed and 25% of patients are diagnosed with meta- Phosphodiesterases static colon cancer. Resistant to chemotherapeutic agents is one of the major problems associated with treating cAMP colon cancer which creates the need to develop novel agents targeting towards newer targets. A phosphodies- cGMP terase is a group of isoenzyme, which, hydrolyze cyclic nucleotides and thereby lowers intracellular levels of Adenylate cyclase cAMP and cGMP leading to tumorigenic effects. Many in vitro and in vivo studies have confirmed increased PDE Guanylate cyclase expression in different types of cancers including colon cancer. cAMP-specific PDE inhibitors increase in- Colon cancer tracellular cAMP that leads to activation of effector molecules-cAMP-dependent protein kinase A, exchange protein activated by cAMP and cAMP gated ion channels. These molecules regulate cellular responses and exert its anticancer role through different mechanisms including apoptosis, inhibition of angiogenesis, upregulating tumor suppressor genes and suppressing oncogenes. On the other hand, cGMP specific PDE inhibitors exhibit anticancer effects through cGMP dependent protein kinase and cGMP dependent cation channels. Elevation in cGMP works through activation of caspases, suppression of Wnt/b-catenin pathway and TCF transcription leading to inhibition of CDK and survivin. These studies point out towards the fact that PDE inhibition is as- sociated with anti-proliferative, anti-apoptotic and anti-angiogenic pathways involved in its anticancer effects in colon cancer.
    [Show full text]
  • Systematic Elucidation of Neuron-Astrocyte Interaction in Models of Amyotrophic Lateral Sclerosis Using Multi-Modal Integrated Bioinformatics Workflow
    ARTICLE https://doi.org/10.1038/s41467-020-19177-y OPEN Systematic elucidation of neuron-astrocyte interaction in models of amyotrophic lateral sclerosis using multi-modal integrated bioinformatics workflow Vartika Mishra et al.# 1234567890():,; Cell-to-cell communications are critical determinants of pathophysiological phenotypes, but methodologies for their systematic elucidation are lacking. Herein, we propose an approach for the Systematic Elucidation and Assessment of Regulatory Cell-to-cell Interaction Net- works (SEARCHIN) to identify ligand-mediated interactions between distinct cellular com- partments. To test this approach, we selected a model of amyotrophic lateral sclerosis (ALS), in which astrocytes expressing mutant superoxide dismutase-1 (mutSOD1) kill wild-type motor neurons (MNs) by an unknown mechanism. Our integrative analysis that combines proteomics and regulatory network analysis infers the interaction between astrocyte-released amyloid precursor protein (APP) and death receptor-6 (DR6) on MNs as the top predicted ligand-receptor pair. The inferred deleterious role of APP and DR6 is confirmed in vitro in models of ALS. Moreover, the DR6 knockdown in MNs of transgenic mutSOD1 mice attenuates the ALS-like phenotype. Our results support the usefulness of integrative, systems biology approach to gain insights into complex neurobiological disease processes as in ALS and posit that the proposed methodology is not restricted to this biological context and could be used in a variety of other non-cell-autonomous communication
    [Show full text]
  • Generation of Mouse Oocytes Defective in Camp Synthesis and Degradation: Endogenous Cyclic AMP Is Essential for Meiotic Arrest
    NIH Public Access Author Manuscript Dev Biol. Author manuscript; available in PMC 2009 October 1. NIH-PA Author ManuscriptPublished NIH-PA Author Manuscript in final edited NIH-PA Author Manuscript form as: Dev Biol. 2008 April 1; 316(1): 124±134. doi:10.1016/j.ydbio.2008.01.018. Generation of mouse oocytes defective in cAMP synthesis and degradation: endogenous cyclic AMP is essential for meiotic arrest Sergio Vaccari1,3, Kathleen Horner1,3, Lisa M. Mehlmann2, and Marco Conti1,3,* 1Division of Reproductive Biology, Department of Obstetrics and Gynecology Stanford University 94305 2Department of Cell Biology, University of Connecticut Health Center, Farmington, CT 06032 Abstract Although it is established that cAMP accumulation plays a pivotal role in preventing meiotic resumption in mammalian oocytes, the mechanisms controlling cAMP levels in the female gamete have remained elusive. Both production of cAMP via GPCRs/Gs/adenylyl cyclases endogenous to the oocyte as well as diffusion from the somatic compartment through gap junctions have been implicated in maintaining cAMP at levels that preclude maturation. Here we have used a genetic approach to investigate the different biochemical pathways contributing to cAMP accumulation and maturation in mouse oocytes. Because cAMP hydrolysis is greatly decreased and cAMP accumulates above a threshold, oocytes deficient in PDE3A do not resume meiosis in vitro or in vivo, resulting in complete female infertility. In vitro, inactivation of Gs or downregulation of the GPCR GPR3 causes meiotic resumption in the Pde3a null oocytes. Crossing of Pde3a-/- mice with Gpr3-/- mice causes partial recovery of female fertility. Unlike the complete meiotic block of the Pde3a null mice, oocyte maturation is restored in the double knockout, although it occurs prematurely as described for the Gpr3-/- mouse.
    [Show full text]
  • Whole-Genome DNA Methylation and Hydroxymethylation Profiling for HBV-Related Hepatocellular Carcinoma
    INTERNATIONAL JOURNAL OF ONCOLOGY 49: 589-602, 2016 Whole-genome DNA methylation and hydroxymethylation profiling for HBV-related hepatocellular carcinoma CHAO YE*, RAN TAO*, QINGYI CAO, DANHUA ZHU, YINI WANG, JIE WANG, JUAN LU, ERMEI CHEN and LANJUAN LI State Key Laboratory for Diagnosis and Treatment of Infectious Diseases, Collaborative Innovation Center for Diagnosis and Treatment of Infectious Diseases, The First Affiliated Hospital, College of Medicine, Zhejiang University, Hangzhou, Zhejiang 310000, P.R. China Received March 18, 2016; Accepted May 13, 2016 DOI: 10.3892/ijo.2016.3535 Abstract. Hepatocellular carcinoma (HCC) is a common tions between them. Taken together, in the present study we solid tumor worldwide with a poor prognosis. Accumulating conducted the first genome-wide mapping of DNA methyla- evidence has implicated important regulatory roles of epigen- tion combined with hydroxymethylation in HBV-related HCC etic modifications in the occurrence and progression of HCC. and provided a series of potential novel epigenetic biomarkers In the present study, we analyzed 5-methylcytosine (5-mC) for HCC. and 5-hydroxymethylcytosine (5-hmC) levels in the tumor tissues and paired adjacent peritumor tissues (APTs) from Introduction four individual HCC patients using a (hydroxy)methylated DNA immunoprecipitation approach combined with deep Hepatocellular carcinoma (HCC), a common solid tumor, is sequencing [(h)MeDIP-Seq]. Bioinformatics analysis revealed the third most frequent cause of cancer-related death in the that the 5-mC levels in the promoter regions of 2796 genes and world. Hepatitis B virus (HBV) infection is the main cause of the 5-hmC levels in 507 genes differed significantly between HCC in China (1).
    [Show full text]
  • G Protein-Regulated Endocytic Trafficking of Adenylyl Cyclase Type 9
    RESEARCH ARTICLE G protein-regulated endocytic trafficking of adenylyl cyclase type 9 Andre´ M Lazar1, Roshanak Irannejad2, Tanya A Baldwin3, Aparna B Sundaram4, J Silvio Gutkind5, Asuka Inoue6, Carmen W Dessauer3, Mark Von Zastrow2,7* 1Program in Biochemistry and Cell Biology, University of California San Francisco, San Francisco, United States; 2Cardiovascular Research Institute and Department of Biochemistry and Biophysics, University of California San Francisco, San Francisco, United States; 3Department of Integrative Biology and Pharmacology, University of Texas Health Science Center, Houston, United States; 4Lung Biology Center, Department of Medicine, University of California San Francisco, San Francisco, United States; 5Department of Pharmacology and Moores Cancer Center, University of California San Diego, San Diego, United States; 6Graduate School of Pharmaceutical Sciences, Tohoku University, Aoba-ku, Sendai, Japan; 7Department of Psychiatry and Department of Cellular and Molecular Pharmacology, University of California San Francisco, San Francisco, United States Abstract GPCRs are increasingly recognized to initiate signaling via heterotrimeric G proteins as they move through the endocytic network, but little is known about how relevant G protein effectors are localized. Here we report selective trafficking of adenylyl cyclase type 9 (AC9) from the plasma membrane to endosomes while adenylyl cyclase type 1 (AC1) remains in the plasma membrane, and stimulation of AC9 trafficking by ligand-induced activation of Gs-coupled GPCRs. AC9 transits a similar, dynamin-dependent early endocytic pathway as ligand-activated GPCRs. However, unlike GPCR traffic control which requires b-arrestin but not Gs, AC9 traffic control requires Gs but not b-arrestin. We also show that AC9, but not AC1, mediates cAMP production stimulated by endogenous receptor activation in endosomes.
    [Show full text]
  • Protein Family Members. the GENE.FAMILY
    Table 3: Protein family members. The GENE.FAMILY col- umn shows the gene family name defined either by HGNC (superscript `H', http://www.genenames.org/cgi-bin/family_ search) or curated manually by us from Entrez IDs in the NCBI database (superscript `C' for `Custom') that we have identified as corresonding for each ENTITY.ID. The members of each gene fam- ily that are in at least one of our synaptic proteome datasets are shown in IN.SYNAPSE, whereas those not found in any datasets are in the column OUT.SYNAPSE. In some cases the intersection of two HGNC gene families are needed to specify the membership of our protein family; this is indicated by concatenation of the names with an ampersand. ENTITY.ID GENE.FAMILY IN.SYNAPSE OUT.SYNAPSE AC Adenylate cyclasesH ADCY1, ADCY2, ADCY10, ADCY4, ADCY3, ADCY5, ADCY7 ADCY6, ADCY8, ADCY9 actin ActinsH ACTA1, ACTA2, ACTB, ACTC1, ACTG1, ACTG2 ACTN ActininsH ACTN1, ACTN2, ACTN3, ACTN4 AKAP A-kinase anchoring ACBD3, AKAP1, AKAP11, AKAP14, proteinsH AKAP10, AKAP12, AKAP17A, AKAP17BP, AKAP13, AKAP2, AKAP3, AKAP4, AKAP5, AKAP6, AKAP8, CBFA2T3, AKAP7, AKAP9, RAB32 ARFGEF2, CMYA5, EZR, MAP2, MYO7A, MYRIP, NBEA, NF2, SPHKAP, SYNM, WASF1 CaM Endogenous ligands & CALM1, CALM2, EF-hand domain CALM3 containingH CaMKK calcium/calmodulin- CAMKK1, CAMKK2 dependent protein kinase kinaseC CB CalbindinC CALB1, CALB2 CK1 Casein kinase 1C CSNK1A1, CSNK1D, CSNK1E, CSNK1G1, CSNK1G2, CSNK1G3 CRHR Corticotropin releasing CRHR1, CRHR2 hormone receptorsH DAGL Diacylglycerol lipaseC DAGLA, DAGLB DGK Diacylglycerol kinasesH DGKB,
    [Show full text]
  • Supplemental Figures 04 12 2017
    Jung et al. 1 SUPPLEMENTAL FIGURES 2 3 Supplemental Figure 1. Clinical relevance of natural product methyltransferases (NPMTs) in brain disorders. (A) 4 Table summarizing characteristics of 11 NPMTs using data derived from the TCGA GBM and Rembrandt datasets for 5 relative expression levels and survival. In addition, published studies of the 11 NPMTs are summarized. (B) The 1 Jung et al. 6 expression levels of 10 NPMTs in glioblastoma versus non‐tumor brain are displayed in a heatmap, ranked by 7 significance and expression levels. *, p<0.05; **, p<0.01; ***, p<0.001. 8 2 Jung et al. 9 10 Supplemental Figure 2. Anatomical distribution of methyltransferase and metabolic signatures within 11 glioblastomas. The Ivy GAP dataset was downloaded and interrogated by histological structure for NNMT, NAMPT, 12 DNMT mRNA expression and selected gene expression signatures. The results are displayed on a heatmap. The 13 sample size of each histological region as indicated on the figure. 14 3 Jung et al. 15 16 Supplemental Figure 3. Altered expression of nicotinamide and nicotinate metabolism‐related enzymes in 17 glioblastoma. (A) Heatmap (fold change of expression) of whole 25 enzymes in the KEGG nicotinate and 18 nicotinamide metabolism gene set were analyzed in indicated glioblastoma expression datasets with Oncomine. 4 Jung et al. 19 Color bar intensity indicates percentile of fold change in glioblastoma relative to normal brain. (B) Nicotinamide and 20 nicotinate and methionine salvage pathways are displayed with the relative expression levels in glioblastoma 21 specimens in the TCGA GBM dataset indicated. 22 5 Jung et al. 23 24 Supplementary Figure 4.
    [Show full text]
  • Detection of H3k4me3 Identifies Neurohiv Signatures, Genomic
    viruses Article Detection of H3K4me3 Identifies NeuroHIV Signatures, Genomic Effects of Methamphetamine and Addiction Pathways in Postmortem HIV+ Brain Specimens that Are Not Amenable to Transcriptome Analysis Liana Basova 1, Alexander Lindsey 1, Anne Marie McGovern 1, Ronald J. Ellis 2 and Maria Cecilia Garibaldi Marcondes 1,* 1 San Diego Biomedical Research Institute, San Diego, CA 92121, USA; [email protected] (L.B.); [email protected] (A.L.); [email protected] (A.M.M.) 2 Departments of Neurosciences and Psychiatry, University of California San Diego, San Diego, CA 92103, USA; [email protected] * Correspondence: [email protected] Abstract: Human postmortem specimens are extremely valuable resources for investigating trans- lational hypotheses. Tissue repositories collect clinically assessed specimens from people with and without HIV, including age, viral load, treatments, substance use patterns and cognitive functions. One challenge is the limited number of specimens suitable for transcriptional studies, mainly due to poor RNA quality resulting from long postmortem intervals. We hypothesized that epigenomic Citation: Basova, L.; Lindsey, A.; signatures would be more stable than RNA for assessing global changes associated with outcomes McGovern, A.M.; Ellis, R.J.; of interest. We found that H3K27Ac or RNA Polymerase (Pol) were not consistently detected by Marcondes, M.C.G. Detection of H3K4me3 Identifies NeuroHIV Chromatin Immunoprecipitation (ChIP), while the enhancer H3K4me3 histone modification was Signatures, Genomic Effects of abundant and stable up to the 72 h postmortem. We tested our ability to use H3K4me3 in human Methamphetamine and Addiction prefrontal cortex from HIV+ individuals meeting criteria for methamphetamine use disorder or not Pathways in Postmortem HIV+ Brain (Meth +/−) which exhibited poor RNA quality and were not suitable for transcriptional profiling.
    [Show full text]
  • ADCY9 on Response to Anacetrapib Among 20,000 Participants in the HPS3/TIMI55-REVEAL Trial
    Impact of ADCY9 on Response to Anacetrapib Among 20,000 Participants in the HPS3/TIMI55-REVEAL Trial Jemma C Hopewell, PhD FESC on behalf of the REVEAL Collaborative Group Disclosures • The REVEAL trial and genetic sub-study was supported by Merck & Co., Inc. • JCH receives no personal payments from industry • JCH is funded by the British Heart Foundation 20 Placebo 15 Background: Dal-OUTCOMES pharmacogenetic study Anacetrapib 10 Randomized 15,871 participants with acute coronary syndrome Participants with Event (%)with Participants 5 Stopped early for futility, and showed no impact of dalcetrapib on cardiovascular outcomes 0 0 1 2 3 4 Years of Follow-up No. at Risk Anacetrapib 15225 14544 13895 13083 8916 Placebo 15224 14545 13896 13029 8746 Absolute benefit 0±2 1±3 5±4 8±4 Schwartz et al, NEJM, 2012 per 1000 participants in anacetrapib group ± SE Source: [P031V01:adam-adpette] Background: Dal-OUTCOMES pharmacogenetic study Dal-OUTCOMES: treatment x genotype interaction: p=0.001 (additive), p=0.006 (genotypic) Tardif et al, Circ CV Gen, 2015 Background: Dal-OUTCOMES pharmacogenetic study Dal-OUTCOMES: treatment x genotype interaction: p=0.001 (additive), p=0.006 (genotypic) The ongoing Dal-genE trial is examining the effects of dalcetrapib in 6150 patients with acute coronary syndrome and the AA genotype Tardif et al, Circ CV Gen, 2015 Background: ACCELERATE pharmacogenetic study ACCELERATE: treatment x genotype interaction: p=0.06 (additive), p=0.17(genotypic) Nissen et al, JAMA Cards 2018 Biological support for an ADCY9 x CETP interaction
    [Show full text]
  • ADCY2, ADCY5, and GRIA1 Are the Key Genes of Camp Signaling Pathway to Participate in Osteoporotic Spinal Fracture After the Manipulation of Wnt Signaling
    ADCY2, ADCY5, and GRIA1 are the key genes of cAMP signaling pathway to participate in osteoporotic spinal fracture after the manipulation of Wnt signaling Type Research paper Keywords Wnt signaling pathway, Spinal Fractures, GRIA1, cAMP signaling Abstract Introduction Osteoporotic spinal fracture, characterized by high morbidity and mortality, has become a health burden for the aging population. The inactivation of the Wnt signaling has been proved to promote osteoporotic fractures. Our study is to identify the key genes, miRNAs, and pathways that possibly lead to osteoporosis and osteoporotic spinal fracture after the aberrant activation or mutation of Wnt signaling pathway. Material and methods Impute R package was used to screen out the differently expressed genes (DEGs) and differently expressed miRNAs in GEO datasets. STRING and Metascape were used to construct protein-protein interactions (PPI) network, gene ontology (GO) enrichment and pathway enrichment. The relative expression of ADCY2, ADCY5, and GRIA1 in bone tissues was measured by RT-qPCR. Results 562 DEGs were screened out using Impute R package, and a PPI network involving the 562 DEGs was constructed using STRING and Metascape. GO enrichment and pathway enrichment showed that the 562 DEGs were associated with membrane protein-related signaling pathways. Then, 75 genes between the target genesPreprint of miR-18a-3p and 562 DEGs were overlapped using Venny 2.1.0. Finally, the cAMP signaling pathway was identified as the key pathway, whilst ADCY2, ADCY5, and GRIA1 were identified the key genes that possibly participate in osteoporotic spinal fracture after the manipulation of Wnt signaling pathway, which was further proved by their excessive downregulation in osteoporotic patients with spinal fracture.
    [Show full text]