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Principal Investigator: Jiexi Chen
Organization: NORTHWESTERN UNIVERSITY AT CHICAGO
Fiscal Year: 2024
Award: $45,336
Funding agency: National Institute of Allergy and Infectious Diseases
Project Summary/Abstract
The marine pathogen Vibrio vulnificus is a highly lethal Gram-negative bacteria inhabiting coastal waters. It
causes intestinal infections via consumption of raw seafood and wound infections via open wounds. The infection
progresses rapidly to septicemia with high fatality rates. This bacterium exerts the highest economic cost of any
seafood-related disease. As multi-antibiotic resistant isolates have been detected and infection incidence is
climbing due to climate change, studies elucidating the pathogenic mechanisms of V. vulnificus are increasingly
important to support novel therapeutic development. The Multifunctional-Autoprocessing Repeats-in-Toxin
(MARTX) toxin has been shown in animal studies to be the primary virulence factor of V. vulnificus associated
with death. The goal of the proposed project is to determine pathogenic mechanisms of MARTX at host cell
surfaces that are necessary for intoxication. Addressing this question could provide critical information toward
design of antibodies that would function to block MARTX from binding to its targeted cells. This goal is particularly
pertinent to the NIAID given the institute's mission to support research to better understand, treat, and prevent
infectious diseases. Specifically, the first aim will determine host receptors for the V. vulnificus MARTXVv toxin.
Our preliminary data using an unbiased Genome-wide CRISPR-Cas9 Screening approach identified a host cell
membrane protein kinase as the most promising receptor candidate. In this aim, we will use a combination of
genetic manipulation, in vitro cellular assays and FRET based flow cytometry to validate the role of this kinase
as a MARTXVv receptor. The second aim will determine which regions of MARTXVv can bind to the cellular surface
based on previous studies on MARTXVv fragments, using techniques including protein purification, cell-based
binding assays and fluorescence microscopy. Collective, these data will define the host receptor and receptor
binding domain of the toxin revealing critical interactions between the toxin and host at cellular surface, the first
step of MARTXVv intoxication. Completion of the proposed work will provide training opportunities required to
support the long-term career goal of becoming a principal investigator studying bacterial toxins. Specifically, the
training will include 1) developing expertise in studying toxin functions in the host cell system, 2) developing
experimental techniques in genetic screens, immunofluorescence, and protein biochemistry and 3) developing
general skills required for academic independent researchers such as grant writing and scientific communication.
Northwestern University Feinberg School of Medicine provides an outstanding training environment for
accomplishing these goals. The sponsor Dr. Karla Satchell is an expert in bacterial toxins and protein structure.
The trainee will gain a depth of knowledge in studying toxin biology by having weekly meetings with her. The
department provides plenty of opportunities for scientific communication including weekly journal club and
research presentations. Further, the program is equipped with excellent core facilities with skillful consultants to
help develop techniques using advanced instruments in imagining, flow cytometry, and protein purification.
Terms: <ATP-protein phosphotransferase><Address><Animals><Antibiotic Resistance><Antibodies><Assay><Bacteria><Bacterial Gene Products><Bacterial Gene Proteins><Bacterial Proteins><Bacterial Toxins><Basic Research><Basic Science><Beneckea><Binding><Bioassay><Biological Assay><Biology><Black Box><Blood Poisoning><C-terminal><CRISPR editing screen><CRISPR screen><CRISPR-based screen><CRISPR/Cas9 screen><Calcium ion><Caspase><Caspase Gene><Cell Body><Cell Communication and Signaling><Cell Death><Cell Line><Cell Membrane Proteins><Cell Signaling><Cell membrane><Cell surface><Cell-Death Protease><CellLine><Cells><Cellular Assay><Cessation of life><Clinical Treatment Moab><Communicable Diseases><Communication><Consensus Sequence><Consumption><Core Facility><Cysteine Endopeptidases><Cysteine Protease><Cysteine Proteinases><Cytolysis><Cytoplasmic Membrane><Cytosol><Cytotoxin><Data><Death><Defect><Disease><Disorder><Environment><Epithelial Cells><Exhibits><FRET><Family><Fatality rate><Flanking Repeat Sequences><Flow Cytofluorometries><Flow Cytofluorometry><Flow Cytometry><Flow Microfluorimetry><Flow Microfluorometry><Fluorescence Light Microscopy><Fluorescence Microscopy><Fluorescence Resonance Energy Transfer><Foundations><Förster Resonance Energy Transfer><Gastroenteritis><Generations><Genes><Genetic Alteration><Genetic Change><Genetic Screening><Genetic defect><Global Warming><Goals><Gram-Negative Bacteria><Grant><Hospital Admission><Hospitalization><Hour><Human><Hydrogen Oxide><ICE-like protease><Immunofluorescence><Immunofluorescence Immunologic><In Vitro><Incidence><Individual><Infection><Infectious Disease Pathway><Infectious Diseases><Infectious Disorder><Inositol Hexakisphosphate><Inositol Hexaphosphate><Intestinal><Intestines><Intoxication><Intracellular Communication and Signaling><Investigators><Journals><Kinase Family Gene><Kinases><Knock-out><Knockout><Knowledge><L-Lysine><Lead><Life><Listonella><Lysine><Lysis><Magazine><Membrane><Membrane Protein Gene><Membrane Proteins><Membrane-Associated Proteins><Microscopic><Mission><Modeling><Modern Man><Molecular Interaction><Monoclonal Antibodies><Multi-Drug Resistance><Multidrug Resistance><Multiple Drug Resistance><Multiple Drug Resistant><Mutation><NIAID><National Institute of Allergy and Infectious Disease><Pathogenesis><Pathogenicity><Pathogenicity Factors><Pb element><Penetration><Phosphotransferase Gene><Phosphotransferases><Phytic Acid><Plasma Membrane><Play><Predisposition><Principal Investigator><Property><Protease Domain><Protein Biochemistry><Protein Fragment><Protein Kinase><Protein/Amino Acid Biochemistry><Proteins><Proteolytic Domain><Public Health><Receptor Protein><Research><Research Personnel><Research Support><Researchers><Resistance><Resistance to Multi-drug><Resistance to Multidrug><Resistance to Multiple Drug><Resistance to antibiotics><Resistant to Multiple Drug><Resistant to antibiotics><Resistant to multi-drug><Resistant to multidrug><Role><Sea-Food><Seafood><Sepsis><Septicemia><Signal Transduction><Signal Transduction Systems><Signaling><Strains Cell Lines><Surface><Surface Proteins><Susceptibility><System><TGF-Beta Type II Receptor><TGFBR2><TGFBR2 gene><Targeted Toxins><Techniques><Terminal Repeat><Terminal Repeat Sequences><Testing><Toxin><Training><Transphosphorylases><Universities><V vulnificus><V. vulnificus><Vibrio><Vibrio vulnificus><Virulence Factors><Water><Work><Wound Infection><Writing><antibiotic drug resistance><antibiotic resistant><biological signal transduction><blood infection><bloodstream infection><bowel><career><cell assay><climate change><climatic changes><clustered regularly interspaced short palindromic repeats screen><coastal water><combat><cultured cell line><cystein protease><cystein proteinase><cysteine endopeptidase><design><designing><economic cost><enteral infection><enteric infection><enteric pathogen infection><enteropathogen infection><enteropathogenic infection><extracellular><flow cytophotometry><food-born><food-born illness><food-borne><food-borne disease><food-borne illness><foodborn><foodborn illness><foodborne><foodborne disease><foodborne illness><gene manipulation><genetic manipulation><genetically manipulate><genetically perturb><genome mutation><genome scale><genome-wide><genomewide><global climate change><glycogen synthase a kinase><gulf coast><heavy metal Pb><heavy metal lead><hydroxyalkyl protein kinase><infected with enteropathogen><infected wound><instrument><intestinal epithelium><intestinal infection><intestine infection><mAbs><marine><marine environment><medical college><medical schools><meeting><meetings><membrane structure><monoclonal Abs><multi-drug resistant><multidrug resistant><necrocytosis><new drug treatments><new drugs><new pharmacological therapeutic><new therapeutics><new therapy><next generation therapeutics><novel drug treatments><novel drugs><novel pharmaco-therapeutic><novel pharmacological therapeutic><novel therapeutics><novel therapy><open wound><pathogen><phosphorylase b kinase kinase><plasmalemma><prevent><preventing><programs><protein purification><protein structure><protein structures><proteins structure><receptor><receptor binding><receptor bound><resistant><school of medicine><septicaemia><septicemic><skills><social role><therapeutic agent development><therapeutic development><tissue wound><toxin V><training opportunity><transforming growth factor-beta type II receptor><transforming growth factor-β type II receptor><wound><wounding><wounds>