Kenneth A. Bradley, Ph.D.

Laboratory Address:
MSB 2817

Mailing Address:
Mail Code: 148906
Los Angeles, CA 90095

Work Address:
MSB 2825
Molecular Sciences Building
Los Angeles, CA 90095 Molecular Sciences Building
Los Angeles, CA 90095

Affiliations
Affiliations
Director, Molecular Screening Shared Resource (MSSR)
Associate Professor, Microbiology, Immunology & Molecular Genetics
Member, Molecular Pharmacology GPB Home Area
Research Interests
The research in our laboratory is inspired by the long-term goal of understanding how microbial pathogens interact with their hosts to promote disease. Specifically, we are interested in studying host genes that are required for bacterial toxins to exert their effects. An example of such a host factor would be a receptor that allows for toxin binding and entry into host cells. We have used anthrax toxin as our model system, but are also beginning to study toxins produced by other pathogenic bacteria such as cholesterol dependent cytolysins (CDCs) and cytolethal distending toxins (CDTs). To achieve this goal, we employ a variety of forward genetic techniques in the host, including somatic cell genetics, chemical genetics, and traditional mouse genetics. Once identified, novel host-pathogen interactions are characterized using biochemical and cell biological strategies to gain a deeper understanding of how these host factors interact with their pathogen-encoded counterparts. ANTHRAX TOXIN Bacillus anthracis produces two major virulence factors, a tripartite exotoxin referred to as anthrax toxin, and an antiphagocytic capsule. These virulence factors mediate pathogen survival and, in the case of the toxin, directly induce damage to the host. Anthrax toxin is an atypical AB toxin in that two distinct catalytic ?A? moieties associate with a single class of binding ?B? moiety. As a result, two separate enzymatic activities are associated with anthrax toxin. The enzymatic subunits are lethal factor (LF), a zinc-dependent metalloproteinase, and edema factor (EF), a calmodulin-dependent adenylate cyclase. LF and EF gain access to the host cytosol via the binding and translocation properties of the shared binding subunit, protective angtigen (PA). Two distinct toxins can be generated by combining either LF or EF with PA. The combination of LF and PA is called lethal toxin (LT), and this toxin inactivates MAPK signaling in the host. Edema toxin, formed by the combination of EF and PA, induces high cAMP levels in host cells. Interestingly, we recently found that ET induces upregulation of anthrax toxin receptors (ANTXRs), thereby inducing a positive feedback loop resulting in increased sensitivity of macrophages and dendritic cells to both LT and ET. Somatic Cell Genetic Approach in Macrophages. Isolation of a receptor-deficient CHO cell line in our original ANTXR cloning efforts {Bradley, 2001} enabled subsequent structure-function studies on ANTXRs. However, macrophages are more likely to be a physiologically relevant target and these cells respond quite differently to LT in culture. Thus, in order to study LT-host cell interactions, we sought to perform somatic cell genetics using a macrophage cell line, RAW264.7. These efforts were successful, and we obtained a macrophage clone that lacks anthrax toxin receptor expression following chemical mutagenesis {Banks, 2005}. This finding demonstrated that forward genetic screening can be accomplished in a macrophage cell line, a cell type which many pathogens have evolved to interact with. In addition, the isolation of the receptor-mutant clone has created opportunities to study the importance of toxin-macrophage interactions during anthrax infection. For example, we were able to show that mutant cells lacking toxin receptors are not only insensitive to purified toxin, but were also able to clear anthrax spores when challenged with a high multiplicity of infection {Banks, 2005; Cote, 2008}. This addressed a long-standing debate about whether toxin production is important during early outgrowth of spores, which occurs within phagolysosomal compartments in the macrophages. ANTXRs were found to co-localize with spores in phagolysosomes, supporting the conclusion that toxin production in this compartment may be important for successful bacterial outgrowth. In collaboration with researches at USAMRIID/Ft. Detrick, we have shown that mice supplemented with macrophages that lack anthrax toxin receptor expression are able to clear a lethal dose of fully virulent B. anthracis spores (Ames strain), while mice supplemented with wild-type or receptor-complemented mutant macrophages cannot {Cote, 2008}. This demonstrated for the first time that toxin targeting of macrophages plays a significant role in vivo for the outcome of an anthrax infection. Development of a conditional, insertion-based somatic cell genetic toolset. While chemical mutagenesis of mammalian cells has proven an effective means to isolate pathogen- and/or toxin-resistant cells {Banks, 2005; Bradley, 2001; Bruce, 2005}, gene identification usually requires cDNA complementation. However, the complementation phenotype is not always easily assayed and/or the gene of interest may not be represented in the cDNA library used. Additionally, with the exception of temperature-sensitive screens, it is not possible to obtain mutant cells that lack a gene essential for cell viability. To overcome these limitations, we developed a novel forward genetic system that allows for conditional, epigenetic control of host gene transcription. We call this system SILENCE for Silencing Induced by Long Terminal Repeat (LTR) Encoded Cis-acting response Element {Banks, 2007}. Chemical Genetics and High Throughput Screening. In addition to somatic cell genetics, we employ chemical genetic approaches to identify host factors involved in anthrax toxin susceptibility. This work is done in conjunction with the Molecular Screening Shared Resource, a high throughput screening (HTS) facility at UCLA. Dr. Bradley currently serves as Director of the MSSR, which has initiated over 150 screening projects for researchers across the United States since 2003. The goal of the chemical screening approach in our lab is two-fold: identify small molecules that can serve as probes to understand toxin action on host cells, and identify small molecules that can serve as lead compounds for development of therapeutics to treat anthrax. To this end, we have reported that two FDA-approved drugs used to treat cardiac arrhythmia or angina block the entry of anthrax toxin at concentrations similar to those found in the serum of human patients {Sanchez, 2007}. These compounds came out of a primary screen of ~500 compounds that represent molecules with known bioactive properties. Based on the initial success of this approach, we expanded our screen to include ~70k small molecules, representing several libraries of diverse chemical structures that are uncharacterized with respect to biological activity. We are currently characterizing a number of hits that resulted from this larger screen. Finally, the HTS capabilities of the MSSR are being utilized to screen genome-wide siRNA and shRNA libraries for genes required for sensitivity to anthrax toxin. CYTOLETHAL DISTENDING TOXINS Cytolethal distending toxins (CDTs) are members of an emerging group of bacterial toxins and effectors called ?cyclomodulins? that interfere with the eukaryotic cell cycle. CDTs intoxicate many cell types and induce G2/M cell cycle arrest, resulting in slow cellular distension and ultimately in cell death. Interfering with the eukaryotic cell cycle has significant implications for pathogens that interact with host cells undergoing rapid division, such as gut epithelium and lymphocytes. CDTs are produced by several unrelated Gram-negative pathogens that induce a wide variety of diseases, including typhoid fever, hemolytic uremic syndrome, and periodontitis. The role of CDT in virulence is poorly understood, but the presence of this toxin results in increased invasiveness, persistence, and/or severity of symptoms. To determine the mechanism by which CDTs interact with mammalian cells, we are undertaking somatic cell genetic and HTS approaches to identify host cell pathways and genes required for efficient toxin uptake and action.
Biography

Dr. Bradley received a B.A. in Biochemistry and Molecular Biology from U.C. Santa Cruz and a Ph.D. in Microbiology and Molecular Genetics from Harvard Medical School. He has been a member of the UCLA faculty since 2002 and is currently an Associate Professor in the Department of Microbiology, Immunology & Molecular Genetics. In addition, Dr. Bradley serves as Director for the Molecular Screening Shared Resource, a high-throughput screening facility that provides robotics for multiple applications including chemical genomics.

Publications

A selected list of publications:

Jung, Michael E; Chamberlain, Brian T; Ho, Chi-Lee C; Gillespie, Eugene J; Bradley, Kenneth A   Structure-Activity Relationship of Semicarbazone EGA Furnishes Photoaffinity Inhibitors of Anthrax Toxin Cellular Entry, ACS Medicinal Chemistry Letters, 2014; .
Gillespie, E. J.; Ho, C. L.; Balaji, K.; Clemens, D. L.; Deng, G.; Wang, Y. E.; Elsaesser, H. J.; Tamilselvam, B.; Gargi, A.; Dixon, S. D.; France, B.; Chamberlain, B. T.; Blanke, S. R.; Cheng, G.; de la Torre, J. C.; Brooks, D. G.; Jung, M. E.; Colicelli, J.; Damoiseaux, R.; Bradley, K. A.   Selective inhibitor of endosomal trafficking pathways exploited by multiple toxins and viruses Proceedings of the National Academy of Sciences of the United States of America, 2013; 110(50): E4904-12.
Gargi, A.; Tamilselvam, B.; Powers, B.; Prouty, M. G.; Lincecum, T.; Eshraghi, A.; Maldonado-Arocho, F. J.; Wilson, B. A.; Bradley, K. A.; Blanke, S. R.   Cellular interactions of the cytolethal distending toxins from Escherichia coli and Haemophilus ducreyi The Journal of biological chemistry, 2013; 288(11): 7492-505.
Ivask, A.; Suarez, E.; Patel, T.; Boren, D.; Ji, Z.; Holden, P.; Telesca, D.; Damoiseaux, R.; Bradley, K. A.; Godwin, H.   Genome-wide bacterial toxicity screening uncovers the mechanisms of toxicity of a cationic polystyrene nanomaterial Environmental science & technology, 2012; 46(4): 2398-405.
Visnyei, K.; Onodera, H.; Damoiseaux, R.; Saigusa, K.; Petrosyan, S.; De Vries, D.; Ferrari, D.; Saxe, J.; Panosyan, E. H.; Masterman-Smith, M.; Mottahedeh, J.; Bradley, K. A.; Huang, J.; Sabatti, C.; Nakano, I.; Kornblum, H. I.   A molecular screening approach to identify and characterize inhibitors of glioblastoma stem cells Molecular cancer therapeutics, 2011; 10(10): 1818-28.
Terra, J. K.; France, B.; Cote, C. K.; Jenkins, A.; Bozue, J. A.; Welkos, S. L.; Bhargava, R.; Ho, C. L.; Mehrabian, M.; Pan, C.; Lusis, A. J.; Davis, R. C.; LeVine, S. M.; Bradley, K. A.   Allelic variation on murine chromosome 11 modifies host inflammatory responses and resistance to Bacillus anthracis PLoS pathogens, 2011; 7(12): e1002469.
Larabee, J. L.; Maldonado-Arocho, F. J.; Pacheco, S.; France, B.; DeGiusti, K.; Shakir, S. M.; Bradley, K. A.; Ballard, J. D.   Glycogen synthase kinase 3 activation is important for anthrax edema toxin-induced dendritic cell maturation and anthrax toxin receptor 2 expression in macrophages Infection and immunity, 2011; 79(8): 3302-8.
Kim, K.; Damoiseaux, R.; Norris, A. J.; Rivina, L.; Bradley, K.; Jung, M. E.; Gatti, R. A.; Schiestl, R. H.; McBride, W. H.   High throughput screening of small molecule libraries for modifiers of radiation responses Int J Radiat Biol, 2011; .
Zhang, H.; Xia, T.; Meng, H.; Xue, M.; George, S.; Ji, Z.; Wang, X.; Liu, R.; Wang, M.; France, B.; Rallo, R.; Damoiseaux, R.; Cohen, Y.; Bradley, K. A.; Zink, J. I.; Nel, A. E.   Differential Expression of Syndecan-1 Mediates Cationic Nanoparticle Toxicity in Undifferentiated versus Differentiated Normal Human Bronchial Epithelial Cells ACS Nano, 2011; .
George, S.; Xia, T.; Rallo, R.; Zhao, Y.; Ji, Z.; Lin, S.; Wang, X.; Zhang, H.; France, B.; Schoenfeld, D.; Damoiseaux, R.; Liu, R.; Bradley, K. A.; Cohen, Y.; Nel, A. E.   Use of a High-Throughput Screening Approach Coupled with In Vivo Zebrafish Embryo Screening To Develop Hazard Ranking for Engineered Nanomaterials ACS Nano, 2011; .
Rallo, R.; France, B.; Liu, R.; Nair, S.; George, S.; Damoiseaux, R.; Giralt, F.; Nel, A.; Bradley, K.; Cohen, Y.   Self-Organizing Map Analysis of Toxicity-Related Cell Signaling Pathways for Metal and Metal Oxide Nanoparticles Environ Sci Technol, 2011; .
Bradley, K. A.; LeVine, S. M.   Anthrax toxin delivers a one-two punch Cell Host Microbe, 2010; 8(5): 394-5.
Ramey, J. D.; Villareal, V. A.; Ng, C.; Ward, S. C.; Xiong, J. P.; Clubb, R. T.; Bradley, K. A.   Anthrax toxin receptor 1/tumor endothelial marker 8: mutation of conserved inserted domain residues overrides cytosolic control of protective antigen binding Biochemistry, 2010; 49(34): 7403-10.
Eshraghi, A.; Maldonado-Arocho, F. J.; Gargi, A.; Cardwell, M. M.; Prouty, M. G.; Blanke, S. R.; Bradley, K. A.   Cytolethal distending toxin family members are differentially affected by alterations in host glycans and membrane cholesterol J Biol Chem, 2010; 285(24): 18199-207.
George, S.; Pokhrel, S.; Xia, T.; Gilbert, B.; Ji, Z.; Schowalter, M.; Rosenauer, A.; Damoiseaux, R.; Bradley, K. A.; Madler, L.; Nel, A. E.   Use of a rapid cytotoxicity screening approach to engineer a safer zinc oxide nanoparticle through iron doping ACS Nano, 2010; 4(1): 15-29.
Terra, J. K.; Cote, C. K.; France, B.; Jenkins, A. L.; Bozue, J. A.; Welkos, S. L.; LeVine, S. M.; Bradley, K. A.   Cutting edge: resistance to Bacillus anthracis infection mediated by a lethal toxin sensitive allele of Nalp1b/Nlrp1b J Immunol, 2010; 184(1): 17-20.
Godwin, H. A.; Chopra, K.; Bradley, K. A.; Cohen, Y.; Harthorn, B. H.; Hoek, E. M.; Holden, P.; Keller, A. A.; Lenihan, H. S.; Nisbet, R. M.; Nel, A. E.   The University of California Center for the Environmental Implications of Nanotechnology Environ Sci Technol, 2009; 43(17): 6453-7.
Maldonado-Arocho, F. J.; Bradley, K. A.   Anthrax edema toxin induces maturation of dendritic cells and enhances chemotaxis towards macrophage inflammatory protein 3beta Infect Immun, 2009; 77(5): 2036-42.
Averette, K. M.; Pratt, M. R.; Yang, Y.; Bassilian, S.; Whitelegge, J. P.; Loo, J. A.; Muir, T. W.; Bradley, K. A.   Anthrax lethal toxin induced lysosomal membrane permeabilization and cytosolic cathepsin release is Nlrp1b/Nalp1b-dependent PLoS ONE, 2009; 4(11): e7913.
Liong, Monty; France, Bryan; Bradley, Kenneth A.; Zink, Jeffrey I.   Antimicrobial Activity of Silver Nanocrystals Encapsulated in Mesoporous Silica Nanoparticles Advanced Materials, 2009; 21(17): 1684-1689.
Lehrer, R. I.; Jung, G.; Ruchala, P.; Wang, W.; Micewicz, E. D.; Waring, A. J.; Gillespie, E. J.; Bradley, K. A.; Ratner, A. J.; Rest, R. F.; Lu, W.   Human alpha-defensins inhibit hemolysis mediated by cholesterol-dependent cytolysins Infect Immun, 2009; 77(9): 4028-40.
Poulos, J. L.; Jeon, T. J.; Damoiseaux, R.; Gillespie, E. J.; Bradley, K. A.; Schmidt, J. J.   Ion channel and toxin measurement using a high throughput lipid membrane platform Biosens Bioelectron, 2009; 24(6): 1806-10.
Lee, S.; Deng, H.; Yu, F.; Melega, W. P.; Damoiseaux, R.; Bradley, K. A.; Sun, R.   Regulation of Kaposi's sarcoma-associated herpesvirus reactivation by dopamine receptor-mediated signaling pathways J Acquir Immune Defic Syndr, 2008; 48(5): 531-40.
Cote, C. K.; DiMezzo, T. L.; Banks, D. J.; France, B.; Bradley, K. A.; Welkos, S. L.   Early interactions between fully virulent Bacillus anthracis and macrophages that influence the balance between spore clearance and development of a lethal infection Microbes Infect, 2008; 10(6): 613-9.
Sanchez, A. M.; Thomas, D.; Gillespie, E. J.; Damoiseaux, R.; Rogers, J.; Saxe, J. P.; Huang, J.; Manchester, M.; Bradley, K. A.   Amiodarone and bepridil inhibit anthrax toxin entry into host cells Antimicrob Agents Chemother, 2007; 51(7): 2403-11.
Nassanian, H.; Sanchez, A. M.; Lo, A.; Bradley, K. A.; Lee, B.   Efficient construction of an inverted minimal H1 promoter driven siRNA expression cassette: facilitation of promoter and siRNA sequence exchange PLoS ONE, 2007; 2(1): e767.
Banks, D. J.; Bradley, K. A.   SILENCE: a new forward genetic technology Nat Methods, 2007; 4(1): 51-3.
Christman, K. L.; Requa, M. V.; Enriquez-Rios, V. D.; Ward, S. C.; Bradley, K. A.; Turner, K. L.; Maynard, H. D.   Submicron streptavidin patterns for protein assembly Langmuir, 2006; 22(17): 7444-50.
Wang, W.; Mulakala, C.; Ward, S. C.; Jung, G.; Luong, H.; Pham, D.; Waring, A. J.; Kaznessis, Y.; Lu, W.; Bradley, K. A.; Lehrer, R. I.   Retrocyclins kill bacilli and germinating spores of Bacillus anthracis and inactivate anthrax lethal toxin J Biol Chem, 2006; .
Maldonado-Arocho, F. J.; Fulcher, J. A.; Lee, B.; Bradley, K. A.   Anthrax oedema toxin induces anthrax toxin receptor expression in monocyte-derived cells Mol Microbiol, 2006; 61(2): 324-37.
Salles,, II; Voth, D. E.; Ward, S. C.; Averette, K. M.; Tweten, R. K.; Bradley, K. A.; Ballard, J. D.   Cytotoxic activity of Bacillus anthracis protective antigen observed in a macrophage cell line overexpressing ANTXR1 Cell Microbiol, 2006; 8(8): 1272-81.
Ding, Z.; Bradley, K. A.; Amin Arnaout, M.; Xiong, J. P.   Expression and purification of functional human anthrax toxin receptor (ATR/TEM8) binding domain from Escherichia coli Protein Expr Purif, 2006; 49(1): 121-8.
Banks, D. J.; Ward, S. C.; Bradley, K. A.   New insights into the functions of anthrax toxin Expert Rev Mol Med, 2006; 8(7): 1-18.
Banks, D. J.; Barnajian, M.; Maldonado-Arocho, F. J.; Sanchez, A. M.; Bradley, K. A.   Anthrax toxin receptor 2 mediates Bacillus anthracis killing of macrophages following spore challenge Cell Microbiol, 2005; 7(8): 1173-85.
Bruce, J. W.; Bradley, K. A.; Ahlquist, P.; Young, J. A.   Isolation of cell lines that show novel, murine leukemia virus-specific blocks to early steps of retroviral replication J Virol, 2005; 79(20): 12969-78.
Sanchez, A. M.; Bradley, K. A.   Anthrax toxin: can a little be a good thing? Trends Microbiol, 2004; 12(4): 143-5.
Bradley, K. A.; Mogridge, J.; Jonah, G.; Rainey, A.; Batty, S.; Young, J. A.   Binding of anthrax toxin to its receptor is similar to alpha integrin-ligand interactions J Biol Chem, 2003; 278(49): 49342-7.
Scobie, H. M.; Rainey, G. J.; Bradley, K. A.; Young, J. A.   Human capillary morphogenesis protein 2 functions as an anthrax toxin receptor Proc Natl Acad Sci U S A, 2003; 100(9): 5170-4.
Bradley, K. A.; Young, J. A.   Anthrax toxin receptor proteins Biochem Pharmacol, 2003; 65(3): 309-14.
Bradley, K. A.; Mogridge, J.; Mourez, M.; Collier, R. J.; Young, J. A.   Identification of the cellular receptor for anthrax toxin Nature, 2001; 414(6860): 225-9.