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Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) is the causative agent of coronavirus disease 2019 (COVID-19), a global pandemic characterized by an exaggerated immune response and respiratory illness. Age (>60 years) is a significant risk factor for developing severe COVID-19. To better understand the host response of the aged airway epithelium to SARS-CoV-2 infection, we performed an in vitro study using primary human bronchial epithelial cells from donors >67 years of age differentiated on an air–liquid interface culture. We demonstrate that SARS-CoV-2 infection leads to early induction of a proinflammatory response and a delayed interferon response. In addition, we observed changes in the genes and pathways associated with cell death and senescence throughout infection. In summary, our study provides new and important insights into the temporal kinetics of the airway epithelial innate immune response to SARS-CoV-2 in older individuals.
Bharathiraja Subramaniyan; Jason L. Larabee; Manish Bodas; Andrew R. Moore; Anthony W. G. Burgett; Dean A. Myers; Constantin Georgescu; Jonathan D. Wren; James F. Papin; Matthew S. Walters. Characterization of the SARS-CoV-2 Host Response in Primary Human Airway Epithelial Cells from Aged Individuals. Viruses 2021, 13, 1603 .
AMA StyleBharathiraja Subramaniyan, Jason L. Larabee, Manish Bodas, Andrew R. Moore, Anthony W. G. Burgett, Dean A. Myers, Constantin Georgescu, Jonathan D. Wren, James F. Papin, Matthew S. Walters. Characterization of the SARS-CoV-2 Host Response in Primary Human Airway Epithelial Cells from Aged Individuals. Viruses. 2021; 13 (8):1603.
Chicago/Turabian StyleBharathiraja Subramaniyan; Jason L. Larabee; Manish Bodas; Andrew R. Moore; Anthony W. G. Burgett; Dean A. Myers; Constantin Georgescu; Jonathan D. Wren; James F. Papin; Matthew S. Walters. 2021. "Characterization of the SARS-CoV-2 Host Response in Primary Human Airway Epithelial Cells from Aged Individuals." Viruses 13, no. 8: 1603.
Clostridioides difficile is a leading cause of nosocomial infection responsible for significant morbidity and mortality with limited options for therapy. Secreted C. difficile toxin B (TcdB) is a major contributor to disease pathology, and select TcdB-specific Abs may protect against disease recurrence. However, the high frequency of recurrence suggests that the memory B cell response, essential for new Ab production following C. difficile reexposure, is insufficient. We therefore isolated TcdB-specific memory B cells from individuals with a history of C. difficile infection and performed single-cell deep sequencing of their Ab genes. Herein, we report that TcdB-specific memory B cell–encoded antibodies showed somatic hypermutation but displayed limited isotype class switch. Memory B cell–encoded mAb generated from the gene sequences revealed low to moderate affinity for TcdB and a limited ability to neutralize TcdB. These findings indicate that memory B cells are an important factor in C. difficile disease recurrence.
Hemangi B. Shah; Kenneth Smith; Edgar J. Scott; Jason L. Larabee; Judith A. James; Jimmy D. Ballard; Mark L. Lang. Human C. difficile toxin–specific memory B cell repertoires encode poorly neutralizing antibodies. JCI Insight 2020, 5, 1 .
AMA StyleHemangi B. Shah, Kenneth Smith, Edgar J. Scott, Jason L. Larabee, Judith A. James, Jimmy D. Ballard, Mark L. Lang. Human C. difficile toxin–specific memory B cell repertoires encode poorly neutralizing antibodies. JCI Insight. 2020; 5 (16):1.
Chicago/Turabian StyleHemangi B. Shah; Kenneth Smith; Edgar J. Scott; Jason L. Larabee; Judith A. James; Jimmy D. Ballard; Mark L. Lang. 2020. "Human C. difficile toxin–specific memory B cell repertoires encode poorly neutralizing antibodies." JCI Insight 5, no. 16: 1.
CREB and C/EBP β signaling pathways are modulated during inflammation and also targeted by Bacillus anthracis edema toxin (ET), but how these factors individually and jointly contribute to changes in immune cell function is poorly understood. Using CRISPR/Cas9 gene editing, macrophage cell lines lacking CREB and isoforms of C/EBP β were generated and analyzed for changes in responses to LPS, ET, and IL-4. Macrophages lacking C/EBP β suppressed induction of IL-10 and Arg1, while IL-6 was increased in these cells following exposure to LPS. Examination of C/EBP β isoforms indicated the 38 kDa isoform was necessary for the expression of IL-10 and Arg1. ChIP-Seq analysis of CREB and C/EBP β binding to targets on the chromosome of human PBMC identified several regions where both factors overlapped in their binding, suggesting similar gene targeting or cooperative effects. Based on the ChIP-Seq data, a panel of previously unknown targets of CREB and C/EBP β was identified and includes genes such as VNN2, GINS4, CTNNBL1, and SULF2. Isoforms of a transcriptional corepressor, transducin-like enhancer of Split (TLE), were also found to have CREB and C/EBP β binding their promoter and were up regulated by ET. Finally, we explore a possible layer of C/EBP β regulation by a protein complex consisting of adenomatous polyposis coli (APC) and PKA. Collectively, these data provide new insights into the role of CREB and C/EBP β as immunosignaling regulators and targets of an important bacterial virulence factor.
Jason L. Larabee; Garrett Hauck; Jimmy D. Ballard. Unique, Intersecting, and Overlapping Roles of C/EBP β and CREB in Cells of the Innate Immune System. Scientific Reports 2018, 8, 16931 .
AMA StyleJason L. Larabee, Garrett Hauck, Jimmy D. Ballard. Unique, Intersecting, and Overlapping Roles of C/EBP β and CREB in Cells of the Innate Immune System. Scientific Reports. 2018; 8 (1):16931.
Chicago/Turabian StyleJason L. Larabee; Garrett Hauck; Jimmy D. Ballard. 2018. "Unique, Intersecting, and Overlapping Roles of C/EBP β and CREB in Cells of the Innate Immune System." Scientific Reports 8, no. 1: 16931.
Clostridium difficile TcdB (2366 amino acid residues) is an intracellular bacterial toxin that binds to cells and enters the cytosol where it glucosylates small GTPases. In the current study, we examined a putative cell entry region of TcdB (amino acid residues 1753-1851) for short sequences that function as cell penetrating peptides (CPPs). To screen for TcdB-derived CPPs, a panel of synthetic peptides were tested for the ability to enhance transferrin (Tf) association with cells. Four candidate CPPs were discovered and further study on one peptide (PepB2) pinpointed an asparagine residue necessary for CPP activity. PepB2 mediated the cell entry of a wide variety of molecules including dextran, streptavidin, microspheres, and lentivirus particles. Of note, this uptake was dramatically reduced in the presence of the Na+/H+ exchange blocker and micropinocytosis inhibitor amiloride, suggesting PepB2 invokes macropinocytosis. Moreover, we found that PepB2 had more efficient cell-penetrating activity than several other well-known CPPs (TAT, Penetratin, Pep-1, or TP10). Finally, Tf assay-based screening of peptides derived from two other large clostridial toxins, TcdA and TcsL, uncovered two new TcdA-derived CPPs. In conclusion, we have identified six CPPs from large clostridial toxins and have demonstrated the ability of PepB2 to promote cell association and entry of several molecules through a putative fluid-phase macropinocytotic mechanism.
Jason L. Larabee; Garrett D. Hauck; Jimmy D. Ballard. Cell-penetrating peptides derived from Clostridium difficile TcdB2 and a related large clostridial toxin. Journal of Biological Chemistry 2018, 293, 1810 -1819.
AMA StyleJason L. Larabee, Garrett D. Hauck, Jimmy D. Ballard. Cell-penetrating peptides derived from Clostridium difficile TcdB2 and a related large clostridial toxin. Journal of Biological Chemistry. 2018; 293 (5):1810-1819.
Chicago/Turabian StyleJason L. Larabee; Garrett D. Hauck; Jimmy D. Ballard. 2018. "Cell-penetrating peptides derived from Clostridium difficile TcdB2 and a related large clostridial toxin." Journal of Biological Chemistry 293, no. 5: 1810-1819.
Edema toxin (ET), composed of edema factor (EF) and protective antigen (PA), is a virulence factor of Bacillus anthracis that alters host immune cell function and contributes to anthrax disease. Anthrax vaccine precipitated (AVP) contains low but detectable levels of EF and can elicit EF-specific antibodies in human recipients of AVP. Active and passive vaccination of mice with EF can contribute to protection from challenge with Bacillus anthracis spores or ET. This study compared humoral responses to ET in recipients of AVP ( n = 33) versus anthrax vaccine adsorbed (AVA; n = 66), matched for number of vaccinations and time postvaccination, and further determined whether EF antibodies elicited by AVP contribute to ET neutralization. AVP induced higher incidence (77.8%) and titer (229.8 ± 58.6) of EF antibodies than AVA (4.2% and 7.8 ± 8.3, respectively), reflecting the reported low but detectable presence of EF in AVP. In contrast, PA IgG levels and ET neutralization measured using a luciferase-based cyclic AMP reporter assay were robust and did not differ between the two vaccine groups. Multiple regression analysis failed to detect an independent contribution of EF antibodies to ET neutralization in AVP recipients; however, EF antibodies purified from AVP sera neutralized ET. Serum samples from at least half of EF IgG-positive AVP recipients bound to nine decapeptides located in EF domains II and III. Although PA antibodies are primarily responsible for ET neutralization in recipients of AVP, increased amounts of an EF component should be investigated for the capacity to enhance next-generation, PA-based vaccines.
Eric K. Dumas; Timothy Gross; Jason Larabee; Lance Pate; Hannah Cuthbertson; Sue Charlton; Bassam Hallis; Renata J. M. Engler; Limone C. Collins; Christina E. Spooner; Hua Chen; Jimmy Ballard; Judith A. James; A. Darise Farris. Anthrax Vaccine Precipitated Induces Edema Toxin-Neutralizing, Edema Factor-Specific Antibodies in Human Recipients. Clinical and Vaccine Immunology 2017, 24, 1 .
AMA StyleEric K. Dumas, Timothy Gross, Jason Larabee, Lance Pate, Hannah Cuthbertson, Sue Charlton, Bassam Hallis, Renata J. M. Engler, Limone C. Collins, Christina E. Spooner, Hua Chen, Jimmy Ballard, Judith A. James, A. Darise Farris. Anthrax Vaccine Precipitated Induces Edema Toxin-Neutralizing, Edema Factor-Specific Antibodies in Human Recipients. Clinical and Vaccine Immunology. 2017; 24 (11):1.
Chicago/Turabian StyleEric K. Dumas; Timothy Gross; Jason Larabee; Lance Pate; Hannah Cuthbertson; Sue Charlton; Bassam Hallis; Renata J. M. Engler; Limone C. Collins; Christina E. Spooner; Hua Chen; Jimmy Ballard; Judith A. James; A. Darise Farris. 2017. "Anthrax Vaccine Precipitated Induces Edema Toxin-Neutralizing, Edema Factor-Specific Antibodies in Human Recipients." Clinical and Vaccine Immunology 24, no. 11: 1.
TcdB is a major virulence factor produced by Clostridium difficile , a leading cause of antibiotic-associated diarrhea. Hypervirulent strains of C. difficile encode a variant of TcdB (TcdB2) that is more toxic than toxin derived from historical strains (TcdB1). Though TcdB1 and TcdB2 exhibit 92% overall identity, a 99-amino-acid region previously associated with cell entry and spanning amino acids 1753 to 1851 has only 77% sequence identity. Results from the present study indicate that the substantial sequence variation in this region could contribute to the differences in cell entry between TcdB1 and TcdB2 and possibly explain TcdB2’s heightened toxicity. Finally, during the course of these studies, an unusual aspect of TcdB cell entry was discovered wherein cell binding appeared to depend on endocytosis. These findings provide insight into TcdB’s variant forms and their mechanisms of cell entry.
Jonathan J. Hunt; Jason L. Larabee; Jimmy D. Ballard. Amino Acid Differences in the 1753-to-1851 Region of TcdB Influence Variations in TcdB1 and TcdB2 Cell Entry. mSphere 2017, 2, e00268-17 .
AMA StyleJonathan J. Hunt, Jason L. Larabee, Jimmy D. Ballard. Amino Acid Differences in the 1753-to-1851 Region of TcdB Influence Variations in TcdB1 and TcdB2 Cell Entry. mSphere. 2017; 2 (4):e00268-17.
Chicago/Turabian StyleJonathan J. Hunt; Jason L. Larabee; Jimmy D. Ballard. 2017. "Amino Acid Differences in the 1753-to-1851 Region of TcdB Influence Variations in TcdB1 and TcdB2 Cell Entry." mSphere 2, no. 4: e00268-17.
Clostridium difficile infection (CDI) is a major cause of hospital-associated, antibiotic-induced diarrhea, which is largely mediated by the production of two large multidomain clostridial toxins, TcdA and TcdB. Both toxins coordinate the action of specific domains to bind receptors, enter cells, and deliver a catalytic fragment into the cytosol. This results in GTPase inactivation, actin disassembly, and cytotoxicity. TcdB in particular has been shown to encode a region covering amino acids 1753 to 1851 that affects epitope exposure and cytotoxicity. Surprisingly, studies here show that several peptides derived from this region, which share the consensus sequence 1769 NVFKGNTISDK 1779 , protect cells from the action of TcdB. One peptide, PepB2, forms multiple interactions with the carboxy-terminal region of TcdB, destabilizes TcdB structure, and disrupts cell binding. We further show that these effects require PepB2 to form a higher-order polymeric complex, a process that requires the central GN amino acid pair. These data suggest that TcdB 1769–1779 interacts with repeat sequences in the proximal carboxy-terminal domain of TcdB (i.e., the CROP domain) to alter the conformation of TcdB. Furthermore, these studies provide insights into TcdB structure and functions that can be exploited to inactivate this critical virulence factor and ameliorate the course of CDI. IMPORTANCE Clostridium difficile is a leading cause of hospital-associated illness that is often associated with antibiotic treatment. To cause disease, C. difficile secretes toxins, including TcdB, which is a multidomain intracellular bacterial toxin that undergoes conformational changes during cellular intoxication. This study describes the development of peptide-based inhibitors that target a region of TcdB thought to be critical for structural integrity of the toxin. The results show that peptides derived from a structurally important region of TcdB can be used to destabilize the toxin and prevent cellular intoxication. Importantly, this work provides a novel means of toxin inhibition that could in the future develop into a C. difficile treatment.
Jason L. Larabee; Sarah J. Bland; Jonathan J. Hunt; Jimmy D. Ballard. Intrinsic Toxin-Derived Peptides Destabilize and Inactivate Clostridium difficile TcdB. mBio 2017, 8, e00503-17 .
AMA StyleJason L. Larabee, Sarah J. Bland, Jonathan J. Hunt, Jimmy D. Ballard. Intrinsic Toxin-Derived Peptides Destabilize and Inactivate Clostridium difficile TcdB. mBio. 2017; 8 (3):e00503-17.
Chicago/Turabian StyleJason L. Larabee; Sarah J. Bland; Jonathan J. Hunt; Jimmy D. Ballard. 2017. "Intrinsic Toxin-Derived Peptides Destabilize and Inactivate Clostridium difficile TcdB." mBio 8, no. 3: e00503-17.
The sequence, activity, and antigenicity of TcdB varies between different strains of Clostridium difficile. As a result, ribotype-specific forms of TcdB exhibit different toxicities and are not strongly cross-neutralized. Using a combination of biochemical and immunological approaches, we compared two important variants of TcdB (TcdB012 and TcdB027) to identify the mechanisms through which sequence differences alter epitopes and activity of the toxin. These analyses led to the discovery of a critical variation in the 1753–1851 (B2′) region of TcdB, which affects the exposure of neutralizing epitopes in the toxin. Sequence comparisons found that the B2′ region exhibits only 77% identity and is the most variable sequence between the two forms of TcdB. A combination of biochemical, analytical, and mutagenesis experiments revealed that the B2′ region promotes protein-protein interactions. These interactions appear to shield neutralizing epitopes that would otherwise be exposed in the toxin, an event found to be less prominent in TcdB012 due to sequence differences in the 1773–1780 and 1791–1798 regions of the B2′ domain. When the carboxyl-terminal domains of TcdB012 and TcdB027 are swapped, neutralization experiments suggest that the amino terminus of TcdB interacts with the B2′ region and impacts the exposure of neutralizing epitopes in the carboxyl terminus. Collectively, these data suggest that variations in the B2′ region affect protein-protein interactions within TcdB and that these interactions influence the exposure of neutralizing epitopes.
Jason Larabee; Aleze Krumholz; Jonathan J. Hunt; Jordi M. Lanis; Jimmy D. Ballard. Exposure of Neutralizing Epitopes in the Carboxyl-terminal Domain of TcdB Is Altered by a Proximal Hypervariable Region. Journal of Biological Chemistry 2015, 290, 6975 -6985.
AMA StyleJason Larabee, Aleze Krumholz, Jonathan J. Hunt, Jordi M. Lanis, Jimmy D. Ballard. Exposure of Neutralizing Epitopes in the Carboxyl-terminal Domain of TcdB Is Altered by a Proximal Hypervariable Region. Journal of Biological Chemistry. 2015; 290 (11):6975-6985.
Chicago/Turabian StyleJason Larabee; Aleze Krumholz; Jonathan J. Hunt; Jordi M. Lanis; Jimmy D. Ballard. 2015. "Exposure of Neutralizing Epitopes in the Carboxyl-terminal Domain of TcdB Is Altered by a Proximal Hypervariable Region." Journal of Biological Chemistry 290, no. 11: 6975-6985.
Here, we describe the capacity ofBacillus anthracispeptidoglycan (BaPGN) to trigger an antimicrobial response in human white blood cells (WBCs). Analysis of freshly isolated human blood cells found that monocytes and neutrophils, but not B and T cells, were highly responsive to BaPGN and produced a variety of cytokines and chemokines. This BaPGN-induced response was suppressed by anthrax lethal toxin (LT) and edema toxin (ET), with the most pronounced effect on human monocytes, and this corresponded with the higher levels of anthrax toxin receptor 1 (ANTXR1) in these cells than in neutrophils. The supernatant from BaPGN-treated cells altered the growth ofB. anthracisSterne, and this effect was blocked by LT, but not by ET. An FtsX mutant ofB. anthracisknown to be resistant to the antimicrobial effects of interferon-inducible Glu-Leu-Arg (ELR)-negative CXC chemokines was not affected by the BaPGN-induced antimicrobial effects. Collectively, these findings describe a system in which BaPGN triggers expression of antimicrobial factors in human WBCs and reveal a distinctive role, not shared with ET, in LT's capacity to suppress this response.
Soumitra Barua; Janaki K. Iyer; Jason L. Larabee; Brent Raisley; Molly A. Hughes; K. Mark Coggeshall; Jimmy D. Ballard. Toxin Inhibition of Antimicrobial Factors Induced by Bacillus anthracis Peptidoglycan in Human Blood. Infection and Immunity 2013, 81, 3693 -3702.
AMA StyleSoumitra Barua, Janaki K. Iyer, Jason L. Larabee, Brent Raisley, Molly A. Hughes, K. Mark Coggeshall, Jimmy D. Ballard. Toxin Inhibition of Antimicrobial Factors Induced by Bacillus anthracis Peptidoglycan in Human Blood. Infection and Immunity. 2013; 81 (10):3693-3702.
Chicago/Turabian StyleSoumitra Barua; Janaki K. Iyer; Jason L. Larabee; Brent Raisley; Molly A. Hughes; K. Mark Coggeshall; Jimmy D. Ballard. 2013. "Toxin Inhibition of Antimicrobial Factors Induced by Bacillus anthracis Peptidoglycan in Human Blood." Infection and Immunity 81, no. 10: 3693-3702.
In cells of the innate immune system, pathological increases in intracellular cAMP attenuate immune responses and contribute to infections by bacteria such as Bacillus anthracis. In this work, cAMP from B. anthracis edema toxin (ET) is found to activate the Notch signaling pathway in both mouse macrophages and human monocytes. ET as well as a cell-permeable activator of PKA induce Notch target genes (HES1, HEY1, IL2RA, and IL7R) and are able to significantly enhance the induction of these Notch target genes by a Toll-like receptor ligand. Elevated cAMP also resulted in increased levels of Groucho/transducin-like enhancer of Split (TLE) and led to increased amounts of a transcriptional repressor complex consisting of TLE and the Notch target Hes1. To address the mechanism used by ET to activate Notch signaling, components of Notch signaling were examined, and results revealed that ET increased levels of recombinant recognition sequence binding protein at the Jκ site (RBP-J), a DNA binding protein and principal transcriptional regulator of Notch signaling. Overexpression studies indicated that RBP-J was sufficient to activate Notch signaling and potentiate LPS-induced Notch signaling. Further examination of the mechanism used by ET to activate Notch signaling revealed that C/EBP β, a transcription factor activated by cAMP, helped activate Notch signaling and up-regulated RBP-J. These studies demonstrate that cAMP activates Notch signaling and increases the expression of TLE, which could be an important mechanism utilized by cAMP to suppress immune responses.
Jason L. Larabee; Salika M. Shakir; Soumitra Barua; Jimmy D. Ballard. Increased cAMP in Monocytes Augments Notch Signaling Mechanisms by Elevating RBP-J and Transducin-like Enhancer of Split (TLE). Journal of Biological Chemistry 2013, 288, 21526 -21536.
AMA StyleJason L. Larabee, Salika M. Shakir, Soumitra Barua, Jimmy D. Ballard. Increased cAMP in Monocytes Augments Notch Signaling Mechanisms by Elevating RBP-J and Transducin-like Enhancer of Split (TLE). Journal of Biological Chemistry. 2013; 288 (30):21526-21536.
Chicago/Turabian StyleJason L. Larabee; Salika M. Shakir; Soumitra Barua; Jimmy D. Ballard. 2013. "Increased cAMP in Monocytes Augments Notch Signaling Mechanisms by Elevating RBP-J and Transducin-like Enhancer of Split (TLE)." Journal of Biological Chemistry 288, no. 30: 21526-21536.
The aim of this study is to determine whether exposure to hazardous chemicals alters chemokine or cytokine production in macrophages and link these events to changes in intracellular signaling pathways and activation of specific gene promoters.
Soumitra Barua; Jason Larabee; James L. Regens; Jimmy D. Ballard. Differential inflammatory responses triggered by toxic small molecules. Environmental Science and Pollution Research 2011, 19, 619 -627.
AMA StyleSoumitra Barua, Jason Larabee, James L. Regens, Jimmy D. Ballard. Differential inflammatory responses triggered by toxic small molecules. Environmental Science and Pollution Research. 2011; 19 (3):619-627.
Chicago/Turabian StyleSoumitra Barua; Jason Larabee; James L. Regens; Jimmy D. Ballard. 2011. "Differential inflammatory responses triggered by toxic small molecules." Environmental Science and Pollution Research 19, no. 3: 619-627.
Anthrax edema toxin (ET) is one of two binary toxins produced by Bacillus anthracis that contributes to the virulence of this pathogen. ET is an adenylate cyclase that generates high levels of cyclic AMP (cAMP), causing alterations in multiple host cell signaling pathways. We previously demonstrated that ET increases cell surface expression of the anthrax toxin receptors (ANTXR) in monocyte-derived cells and promotes dendritic cell (DC) migration toward the lymph node-homing chemokine MIP-3β. In this work, we sought to determine if glycogen synthase kinase 3 (GSK-3) is important for ET-induced modulation of macrophage and DC function. We demonstrate that inhibition of GSK-3 dampens ET-induced maturation and migration processes of monocyte-derived dendritic cells (MDDCs). Additional studies reveal that the ET-induced expression of ANTXR in macrophages was decreased when GSK-3 activity was disrupted with chemical inhibitors or with small interfering RNA (siRNA) targeting GSK-3. Further examination of the ET induction of ANTXR revealed that a dominant negative form of CREB could block the ET induction of ANTXR, suggesting that CREB or a related family member was involved in the upregulation of ANTXR. Because CREB and GSK-3 activity appeared to be important for ET-induced ANTXR expression, the impact of GSK-3 on ET-induced CREB activity was examined in RAW 264.7 cells possessing a CRE-luciferase reporter. As with ANTXR expression, the ET induction of the CRE reporter was decreased by reducing GSK-3 activity. These studies not only provide insight into host pathways targeted by ET but also shed light on interactions between GSK-3 and CREB pathways in host immune cells.
Jason L. Larabee; Francisco J. Maldonado-Arocho; Sergio Pacheco; Bryan France; Kevin DeGiusti; Salika M. Shakir; Kenneth A. Bradley; Jimmy D. Ballard. 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, 3302 -3308.
AMA StyleJason L. Larabee, Francisco J. Maldonado-Arocho, Sergio Pacheco, Bryan France, Kevin DeGiusti, Salika M. Shakir, Kenneth A. Bradley, Jimmy D. Ballard. 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):3302-3308.
Chicago/Turabian StyleJason L. Larabee; Francisco J. Maldonado-Arocho; Sergio Pacheco; Bryan France; Kevin DeGiusti; Salika M. Shakir; Kenneth A. Bradley; Jimmy D. Ballard. 2011. "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 79, no. 8: 3302-3308.
The production of cAMP from Bacillus anthracis edema toxin (ET) activates gene expression in macrophages through a complex array of signaling pathways, most of which remain poorly defined. In this study, the tumor suppressor protein adenomatous polyposis coli (APC) was found to be important for the up-regulation of previously defined ET-stimulated genes (Vegfa, Ptgs2, Arg2, Cxcl2, Sdc1, and Cebpb). A reduction in the expression of these genes after ET exposure was observed when APC was disrupted in macrophages using siRNA or in bone marrow-derived macrophages obtained from C57BL/6J-Apc(Min) mice, which are heterozygous for a truncated form of APC. In line with this observation, ET increased the expression of APC at the transcriptional level, leading to increased amounts of APC in the nucleus. The mechanism utilized by APC to increase ET-induced gene expression was determined to depend on the ability of APC to interact with C/EBP β, which is a transcription factor activated by cAMP. Coimmunoprecipitation experiments found that APC associated with C/EBP β and that levels of this complex increase after ET exposure. A further connection was uncovered when silencing APC was determined to reduce the ET-induced phosphorylation of C/EBP β at Thr-188. This ET-mediated phosphorylation of C/EBP β was blocked by glycogen synthase kinase 3 (GSK-3) inhibitors, suggesting that GSK-3 is involved in the activation of C/EBP β and supporting the idea of APC helping direct interactions between GSK-3 and C/EBP β. These results indicate that ET stimulates gene expression by promoting the formation of an inducible protein complex consisting of APC and C/EBP β.
Jason L. Larabee; Salika M. Shakir; Logan Hightower; Jimmy D. Ballard. Adenomatous Polyposis Coli Protein Associates with C/EBP β and Increases Bacillus anthracis Edema Toxin-stimulated Gene Expression in Macrophages. Journal of Biological Chemistry 2011, 286, 19364 -19372.
AMA StyleJason L. Larabee, Salika M. Shakir, Logan Hightower, Jimmy D. Ballard. Adenomatous Polyposis Coli Protein Associates with C/EBP β and Increases Bacillus anthracis Edema Toxin-stimulated Gene Expression in Macrophages. Journal of Biological Chemistry. 2011; 286 (22):19364-19372.
Chicago/Turabian StyleJason L. Larabee; Salika M. Shakir; Logan Hightower; Jimmy D. Ballard. 2011. "Adenomatous Polyposis Coli Protein Associates with C/EBP β and Increases Bacillus anthracis Edema Toxin-stimulated Gene Expression in Macrophages." Journal of Biological Chemistry 286, no. 22: 19364-19372.
Limited information is available regarding systemic changes in mammals associated with exposures to petroleum/hydrocarbon fuels. In this study, systemic toxicity of JP-8 jet fuel was observed in a rat inhalation model at different JP-8 fuel vapor concentrations (250, 500, or 1000 mg/m(3), for 91 days). Gel electrophoresis and mass spectrometry sequencing identified the alpha-2 microglobulin protein to be elevated in rat kidney in a JP-8 dose-dependent manner. Western blot analysis of kidney and lung tissue extracts revealed JP-8 dependent elevation of inducible heat shock protein 70 (HSP70). Tissue changes were observed histologically (hematoxylin and eosin staining) in liver, kidney, lung, bone marrow, and heart, and more prevalently at medium or high JP-8 vapor phase exposures (500-1000 mg/m(3)) than at low vapor phase exposure (250 mg/m(3)) or non-JP-8 controls. JP-8 fuel-induced liver alterations included dilated sinusoids, cytoplasmic clumping, and fat cell deposition. Changes to the kidneys included reduced numbers of nuclei, and cytoplasmic dumping in the lumen of proximal convoluted tubules. JP-8 dependent lung alterations were edema and dilated alveolar capillaries, which allowed clumping of red blood cells (RBCs). Changes in the bone marrow in response to JP-8 included reduction of fat cells and fat globules, and cellular proliferation (RBCs, white blood cells-WBCs, and megakaryocytes). Heart tissue from JP-8 exposed animals contained increased numbers of inflammatory and fibroblast cells, as well as myofibril scarring. cDNA array analysis of heart tissue revealed a JP-8 dependent increase in atrial natriuretic peptide precursor mRNA and a decrease in voltage-gated potassium (K+) ion channel mRNA.
Jay S. Hanas; G. Bruce Briggs; Megan R. Lerner; Stan A. Lightfoot; Jason L. Larabee; Todd J. Karsies; Robert B. Epstein; Rushie J. Hanas; Daniel J. Brackett; James R. Hocker. Systemic molecular and cellular changes induced in rats upon inhalation of JP-8 petroleum fuel vapor. Toxicology Mechanisms and Methods 2010, 20, 204 -212.
AMA StyleJay S. Hanas, G. Bruce Briggs, Megan R. Lerner, Stan A. Lightfoot, Jason L. Larabee, Todd J. Karsies, Robert B. Epstein, Rushie J. Hanas, Daniel J. Brackett, James R. Hocker. Systemic molecular and cellular changes induced in rats upon inhalation of JP-8 petroleum fuel vapor. Toxicology Mechanisms and Methods. 2010; 20 (4):204-212.
Chicago/Turabian StyleJay S. Hanas; G. Bruce Briggs; Megan R. Lerner; Stan A. Lightfoot; Jason L. Larabee; Todd J. Karsies; Robert B. Epstein; Rushie J. Hanas; Daniel J. Brackett; James R. Hocker. 2010. "Systemic molecular and cellular changes induced in rats upon inhalation of JP-8 petroleum fuel vapor." Toxicology Mechanisms and Methods 20, no. 4: 204-212.
In the current study, we examined the regulatory interactions of a serine/threonine phosphatase (BA-Stp1), serine/threonine kinase (BA-Stk1) pair in Bacillus anthracis . B. anthracis STPK101, a null mutant lacking BA-Stp1 and BA-Stk1, was impaired in its ability to survive within macrophages, and this correlated with an observed reduction in virulence in a mouse model of pulmonary anthrax. Biochemical analyses confirmed that BA-Stp1 is a PP2C phosphatase and dephosphorylates phosphoserine and phosphothreonine residues. Treatment of BA-Stk1 with BA-Stp1 altered BA-Stk1 kinase activity, indicating that the enzymatic function of BA-Stk1 can be influenced by BA-Stp1 dephosphorylation. Using a combination of mass spectrometry and mutagenesis approaches, three phosphorylated residues, T165, S173, and S214, in BA-Stk1 were identified as putative regulatory targets of BA-Stp1. Further analysis found that T165 and S173 were necessary for optimal substrate phosphorylation, while S214 was necessary for complete ATP hydrolysis, autophosphorylation, and substrate phosphorylation. These findings provide insight into a previously undescribed Stp/Stk pair in B. anthracis .
Salika M. Shakir; Katie M. Bryant; Jason L. Larabee; Elaine E. Hamm; Julie Lovchik; C. Rick Lyons; Jimmy D. Ballard. Regulatory Interactions of a Virulence-Associated Serine/Threonine Phosphatase-Kinase Pair in Bacillus anthracis. Journal of Bacteriology 2010, 192, 400 -409.
AMA StyleSalika M. Shakir, Katie M. Bryant, Jason L. Larabee, Elaine E. Hamm, Julie Lovchik, C. Rick Lyons, Jimmy D. Ballard. Regulatory Interactions of a Virulence-Associated Serine/Threonine Phosphatase-Kinase Pair in Bacillus anthracis. Journal of Bacteriology. 2010; 192 (2):400-409.
Chicago/Turabian StyleSalika M. Shakir; Katie M. Bryant; Jason L. Larabee; Elaine E. Hamm; Julie Lovchik; C. Rick Lyons; Jimmy D. Ballard. 2010. "Regulatory Interactions of a Virulence-Associated Serine/Threonine Phosphatase-Kinase Pair in Bacillus anthracis." Journal of Bacteriology 192, no. 2: 400-409.
Exogenous CD1d-binding glycolipid (alpha-Galactosylceramide, alpha-GC) stimulates TCR signaling and activation of type-1 natural killer-like T (NKT) cells. Activated NKT cells play a central role in the regulation of adaptive and protective immune responses against pathogens and tumors. In the present study, we tested the effect of Bacillus anthracis lethal toxin (LT) on NKT cells both in vivo and in vitro. LT is a binary toxin known to suppress host immune responses during anthrax disease and intoxicates cells by protective antigen (PA)-mediated intracellular delivery of lethal factor (LF), a potent metalloprotease. We observed that NKT cells expressed anthrax toxin receptors (CMG-2 and TEM-8) and bound more PA than other immune cell types. A sub-lethal dose of LT administered in vivo in C57BL/6 mice decreased expression of the activation receptor NKG2D by NKT cells but not by NK cells. The in vivo administration of LT led to decreased TCR-induced cytokine secretion but did not affect TCR expression. Further analysis revealed LT-dependent inhibition of TCR-stimulated MAP kinase signaling in NKT cells attributable to LT cleavage of the MAP kinase kinase MEK-2. We propose that Bacillus anthracis-derived LT causes a novel form of functional anergy in NKT cells and therefore has potential for contributing to immune evasion by the pathogen
Sunil K. Joshi; Gillian A. Lang; Jason L. Larabee; T. Scott Devera; Lindsay M. Aye; Hemangi B. Shah; Jimmy D. Ballard; Mark L. Lang. Bacillus anthracis Lethal Toxin Disrupts TCR Signaling in CD1d-Restricted NKT Cells Leading to Functional Anergy. PLOS Pathogens 2009, 5, e1000588 .
AMA StyleSunil K. Joshi, Gillian A. Lang, Jason L. Larabee, T. Scott Devera, Lindsay M. Aye, Hemangi B. Shah, Jimmy D. Ballard, Mark L. Lang. Bacillus anthracis Lethal Toxin Disrupts TCR Signaling in CD1d-Restricted NKT Cells Leading to Functional Anergy. PLOS Pathogens. 2009; 5 (9):e1000588.
Chicago/Turabian StyleSunil K. Joshi; Gillian A. Lang; Jason L. Larabee; T. Scott Devera; Lindsay M. Aye; Hemangi B. Shah; Jimmy D. Ballard; Mark L. Lang. 2009. "Bacillus anthracis Lethal Toxin Disrupts TCR Signaling in CD1d-Restricted NKT Cells Leading to Functional Anergy." PLOS Pathogens 5, no. 9: e1000588.
The anti-inflammatory selenium compounds, ebselen (2-phenyl-1,2-benzisoselenazol-3[2H]-one) and selenite, were found to alter the DNA binding mechanisms and structures of cysteine-rich zinc-finger transcription factors. As assayed by DNase I protection, DNA binding by TFIIIA (transcription factor IIIA, prototypical Cys2His2 zinc finger protein), was inhibited by micromolar amounts of ebselen. In a gel shift assay, ebselen inhibited the Cys2His2 zinc finger-containing DNA binding domain (DBD) of the NF-κB mediated transcription factor Sp1. Ebselen also inhibited DNA binding by the p50 subunit of the pro-inflammatory Cys-containing NF-κB transcription factor. Electrospray ionization mass spectrometry (ESI-MS) was utilized to elucidate mechanisms of chemical interaction between ebselen and a zinc-bound Cys2His2 zinc finger polypeptide modeled after the third finger of Sp1 (Sp1-3). Exposing Sp1-3 to micromolar amounts of ebselen resulted in Zn2+ release from this peptide and the formation of a disulfide bond by oxidation of zinc finger SH groups, the likely mechanism for DNA binding inhibition. Selenite was shown by ESI-MS to also eject zinc from Sp1-3 as well as induce disulfide bond formation through SH oxidation. The selenite-dependent inhibition/oxidation mechanism differed from that of ebselen by inducing the formation of a stable selenotrisulfide bond. Selenite-induced selenotrisulfide formation was dependent upon the structure of the Cys2His2 zinc finger as alteration in the finger structure enhanced this reaction as well as selenite-dependent zinc release. Ebselen and selenite-dependent inhibition/oxidation of Cys-rich zinc finger proteins, with concomitant release of zinc and finger structural changes, points to mechanisms at the atomic and protein level for selenium-induced alterations in Cys-rich proteins, and possible amelioration of certain inflammatory, neurodegenerative, and oncogenic responses.
Jason Larabee; James Hocker; Jay S. Hanas. Mechanisms of inhibition of zinc-finger transcription factors by selenium compounds ebselen and selenite. Journal of Inorganic Biochemistry 2009, 103, 419 -426.
AMA StyleJason Larabee, James Hocker, Jay S. Hanas. Mechanisms of inhibition of zinc-finger transcription factors by selenium compounds ebselen and selenite. Journal of Inorganic Biochemistry. 2009; 103 (3):419-426.
Chicago/Turabian StyleJason Larabee; James Hocker; Jay S. Hanas. 2009. "Mechanisms of inhibition of zinc-finger transcription factors by selenium compounds ebselen and selenite." Journal of Inorganic Biochemistry 103, no. 3: 419-426.
In an effort to better understand the mechanisms by which Bacillus anthracis establishes disease, experiments were undertaken to identify the genes essential for intracellular germination. Eighteen diverse genetic loci were identified via an enrichment protocol using a transposon-mutated library of B. anthracis spores, which was screened for mutants delayed in intracellular germination. Fourteen transposon mutants were identified in genes not previously associated with B. anthracis germination and included disruption of factors involved in membrane transport, transcriptional regulation, and intracellular signaling. Four mutants contained transposon insertions in gerHA, gerHB, gerHC , and pagA , respectively, each of which has been previously associated with germination or survival of B. anthracis within macrophages. Strain MIGD101 (named for macrophage intracellular germination defective 101) was of particular interest, since this mutant contained a transposon insertion in an intergenic region between BAs2807 and BAs2808, and was the most highly represented mutant in the enrichment. Analysis of B. anthracis MIGD101 by confocal microscopy and differential heat sensitivity following macrophage infection revealed ungerminated spores within the cell. Moreover, B. anthracis MIGD101 was attenuated in cell killing relative to the parent strain. Further experimental analysis found that B. anthracis MIGD101 was defective in five known B. anthracis germination pathways, supporting a mechanism wherein the intergenic region between BAs2807 and BAs2808 has a global affect on germination of this pathogen. Collectively, these findings provide insight into the mechanisms supporting B. anthracis germination within host cells.
Soumitra Barua; Matthew McKevitt; Kevin DeGiusti; Elaine E. Hamm; Jason Larabee; Salika Shakir; Katie Bryant; Theresa M. Koehler; Steven R. Blanke; David Dyer; Allison Gillaspy; Jimmy D. Ballard. The Mechanism of Bacillus anthracis Intracellular Germination Requires Multiple and Highly Diverse Genetic Loci. Infection and Immunity 2009, 77, 23 -31.
AMA StyleSoumitra Barua, Matthew McKevitt, Kevin DeGiusti, Elaine E. Hamm, Jason Larabee, Salika Shakir, Katie Bryant, Theresa M. Koehler, Steven R. Blanke, David Dyer, Allison Gillaspy, Jimmy D. Ballard. The Mechanism of Bacillus anthracis Intracellular Germination Requires Multiple and Highly Diverse Genetic Loci. Infection and Immunity. 2009; 77 (1):23-31.
Chicago/Turabian StyleSoumitra Barua; Matthew McKevitt; Kevin DeGiusti; Elaine E. Hamm; Jason Larabee; Salika Shakir; Katie Bryant; Theresa M. Koehler; Steven R. Blanke; David Dyer; Allison Gillaspy; Jimmy D. Ballard. 2009. "The Mechanism of Bacillus anthracis Intracellular Germination Requires Multiple and Highly Diverse Genetic Loci." Infection and Immunity 77, no. 1: 23-31.
Bacillus anthracis edema toxin (ET) generates high levels of cyclic AMP and impacts a complex network of signaling pathways in targeted cells. In the current study, we sought to identify kinase signaling pathways modulated by ET to better understand how this toxin alters cell physiology. Using a panel of small-molecule inhibitors of mammalian kinases, we found that inhibitors of glycogen synthase kinase 3 beta (GSK-3β) protected cells from ET-induced changes in the cell cycle. GSK-3β inhibitors prevented declines in cellular levels of cyclin D1 and c-Jun following treatment of macrophages with ET. Strikingly, cell fractionation experiments and confocal immunofluorescence microscopy revealed that ET activates a compartmentalized pool of GSK-3β residing in the nuclei, but not in the cytoplasm, of macrophages. To investigate the outcome of this event, we examined the cellular location and activation state of β-catenin, a critical substrate of GSK-3β, and found that the protein was inactivated within the nucleus following intoxication with ET. To determine if ET could overcome the effects of stimuli that inactivate GSK-3β, we examined the impact of the toxin on the Wnt signaling pathway. The results of these experiments revealed that by targeting GSK-3β residing in the nucleus, ET circumvents the upstream cytoplasmic inactivation of GSK-3β, which occurs following exposure to Wnt-3A. These findings suggest ET arrests the cell cycle by a mechanism involving activation of GSK-3β residing in the nucleus, and by using this novel mechanism of intoxication, ET avoids cellular systems that would otherwise reverse the effects of the toxin.
Jason L. Larabee; Kevin DeGiusti; James L. Regens; Jimmy D. Ballard. Bacillus anthracis Edema Toxin Activates Nuclear Glycogen Synthase Kinase 3β. Infection and Immunity 2008, 76, 4895 -4904.
AMA StyleJason L. Larabee, Kevin DeGiusti, James L. Regens, Jimmy D. Ballard. Bacillus anthracis Edema Toxin Activates Nuclear Glycogen Synthase Kinase 3β. Infection and Immunity. 2008; 76 (11):4895-4904.
Chicago/Turabian StyleJason L. Larabee; Kevin DeGiusti; James L. Regens; Jimmy D. Ballard. 2008. "Bacillus anthracis Edema Toxin Activates Nuclear Glycogen Synthase Kinase 3β." Infection and Immunity 76, no. 11: 4895-4904.
Dermal exposure to JP-8 petroleum jet fuel leads to toxicological responses in humans and rodents. Serum profiling is a molecular analysis of changes in the levels of serum proteins and other molecules in response to changes in physiology. This present study utilizes serum profiling approaches to examine biomolecular changes in the sera of rats exposed to dermal applications of JP-8 (jet propulsion fuel-8). Using gel electrophoresis and electrospray ionization (ESI) mass spectrometry (MS), levels of serum proteins as well as low-mass constituents were found to change after dermal exposures to JP-8. The serum protein levels altered included the acute-phase response proteins haptoglobin, ceruloplasmin, α1-inhibitor III, and apolipoprotein A-IV. Haptoglobin levels increased after a 1-day JP-8 dermal exposure and continued to increase through 7 days of exposure. Ceruloplasmin levels increased after 5 days of exposure. Serum α1-inhibitor III was reduced after a 1-day exposure and the depletion continued after 7 days of exposure. Apolipoprotein A-IV increased after a 1-day exposure and then returned to basal levels after 3- and 5-day exposures of JP-8. Levels of the acute-phase protein α2-macroglobulin were found to not vary over these time course studies. Using ESI-MS analysis directly on the sera from rats exposed to dermal JP-8, low-mass sera constituents were found to correlate with control (acetone) or JP-8 exposure.
Jason L. Larabee; James R. Hocker; John Y. Cheung; Randle M. Gallucci; Jay S. Hanas. Serum Profiling of Rat Dermal Exposure to JP-8 Fuel Reveals an Acute-Phase Response. Toxicology Mechanisms and Methods 2008, 18, 41 -51.
AMA StyleJason L. Larabee, James R. Hocker, John Y. Cheung, Randle M. Gallucci, Jay S. Hanas. Serum Profiling of Rat Dermal Exposure to JP-8 Fuel Reveals an Acute-Phase Response. Toxicology Mechanisms and Methods. 2008; 18 (1):41-51.
Chicago/Turabian StyleJason L. Larabee; James R. Hocker; John Y. Cheung; Randle M. Gallucci; Jay S. Hanas. 2008. "Serum Profiling of Rat Dermal Exposure to JP-8 Fuel Reveals an Acute-Phase Response." Toxicology Mechanisms and Methods 18, no. 1: 41-51.