bluuebunny/crossref_metadata_2025_split huggingface dataset:
Innate Protection of
Mycobacterium smegmatis
against the Antimicrobial Activity of Nitric Oxide Is Provided by Mycothiol
Authors: Miller Christopher C. , Rawat Mamta , Johnson Todd , Av-Gay Yossef 
Published: July 2007

ABSTRACT

Nitric oxide (NO) is an efficient antimicrobial agent. A role for mycothiol in protecting mycobacteria from nitrosative damage was revealed by showing that a
Mycobacterium smegmatis
mutant is sensitive to NO. A direct correlation between NO and mycothiol levels confirmed that mycothiol is important for protecting mycobacteria from NO attack.
Functional Consequences of Sequence Variation in Bundlin, the Enteropathogenic
Escherichia coli
Type IV Pilin Protein
Authors: Fernandes Paula J. , Guo Qin , Donnenberg Michael S. 
Published: July 2007

ABSTRACT

The bundle-forming pilus (BFP) of enteropathogenic
Escherichia coli
(EPEC) is an important virulence factor. We examined the role of divergent alleles of
bfpA
encoding bundlin, the BFP pilin protein, in pilus biogenesis, pilus interactions, and immune responses. We found that the BFP biogenesis machine from an EPEC strain that expresses one bundlin type is capable of assembling all other bundlin types. Furthermore, we found that EPEC strains expressing divergent bundlin types are capable of forming mixed autoaggregates, suggesting that different pilin types can intertwine. However, we found that there was a marked difference between alleles in immunogenicity in both rabbits and mice of a peptide derived from a region of bundlin undergoing apparent diversifying selection. In addition, despite a high degree of cross-reactivity between divergent bundlin proteins, in both mice and rabbits responses appeared to be stronger against the homologous pilin protein than against the heterologous protein. This result was verified using sera from a volunteer study, which demonstrated that the human antibody responses after an initial challenge with live EPEC were stronger against the homologous bundlin protein than against a divergent bundlin protein. However, a repeat challenge induced equivalent responses. These results are consistent with the hypothesis that human immune responses against bundlin exert selective pressure on
bfpA
sequence divergence.
Differential CD28 and Inducible Costimulatory Molecule Signaling Requirements for Protective CD4+T-Cell-Mediated Immunity against Genital TractChlamydia trachomatisInfection
Authors: Marks Ellen , Verolin Martina , Stensson Anneli , Lycke Nils 
Published: July 2007

ABSTRACTTh1 cells and gamma interferon (IFN-γ) production play critical roles in protective immunity against genital tract infections byChlamydia trachomatis. Here we show that inducible costimulatory molecule (ICOS)−/−mice develop greatly augmented host resistance against chlamydial infection. Protection following a primary infection was characterized by strong Th1 immunity with enhanced CD4+T-cell-mediated IFN-γ production in the genital tract and high expression of T-bet in the draining para-aortic lymph node. This Th1 dominance was associated with low expression of interleukin 10 (IL-10) mRNA in the uteruses of protected ICOS−/−mice. By contrast, CD28−/−mice were severely impaired in their adaptive immune response, demonstrating a lack of CD4+T cells and IFN-γ in the genital tract, with a substantial delay in bacterial elimination compared to that seen in wild-type (WT) mice. Upon reinfection, WT mice exhibited a transient local infection with evidence of regulatory T-cell (Treg)/Foxp3 mRNA and a more balanced Th1 and Th2 response in the genital tract than ICOS−/−mice, whereas 90% of the latter mice developed sterile immunity, poor expression of local Treg/Foxp3 mRNA, and macroscopic signs of enhanced local immunopathology. Therefore, different requirements for CD28 signaling and ICOS signaling clearly apply to host protection against a genital tract infection byC. trachomatis. Whereas, CD28 signaling is critical, ICOS appears to be dispensable and can have a dampening effect on Th1 development by driving Th2 immunity and anti-inflammation through IL-10 production and promotion of the Foxp3+Treg populations in the genital tract. Both the CD28-deficient and the ICOS-deficient mice demonstrated poor specific antibody production, supporting the fact that antibodies are not needed for protection against genital tract chlamydial infections.
Cross-Linked Forms of the Isolated N-Terminal Domain of the Lethal Factor Are Potent Inhibitors of Anthrax Toxin
Authors: Juris Stephen J. , Melnyk Roman A. , Bolcome Robert E. , Chan Joanne , Collier R. John 
Published: July 2007

ABSTRACT

The proteins that comprise anthrax toxin self-assemble at the mammalian cell surface into a series of toxic complexes, each containing a heptameric form of protective antigen (PA) plus up to a total of three molecules of the enzymatic moieties of the toxin (lethal factor [LF] and edema factor [EF]). These complexes are trafficked to the endosome, where the PA heptamer forms a pore in the membrane under the influence of low pH, and bound LF and EF unfold and translocate through the pore to the cytosol. To explore the hypothesis that the PA pore can translocate multiple, cross-linked polypeptides simultaneously, we cross-linked LF
N
, the N-terminal domain of LF, via an introduced cysteine at its N or C terminus and characterized the products. Both dimers and trimers of LF
N
retained the ability to bind to PA pores and block ion conductance, but they were unable to translocate across the membrane, even at high voltages or with a transmembrane pH gradient. The multimers were remarkably potent inhibitors of toxin action in mammalian cells (20- to 50-fold more potent than monomeric LF
N
) and in a zebrafish model system. These findings show that the PA pore cannot translocate multimeric, cross-linked polypeptides and demonstrate a new approach to generating potent inhibitors of anthrax toxin.
Human Defensin α-1 CausesTrypanosoma cruziMembrane Pore Formation and Induces DNA Fragmentation, Which Leads to Trypanosome Destruction
Authors: Madison M. Nia , Kleshchenko Yuliya Y. , Nde Pius N. , Simmons Kaneatra J. , Lima Maria F. , Villalta Fernando 
Published: July 2007

ABSTRACTHuman defensins play a fundamental role in the initiation of innate immune responses to some microbial pathogens. Here we show that human defensin α-1 displays a trypanocidal role againstTrypanosoma cruzi, the causative agent of Chagas' disease. The toxicity of human defensin α-1 againstT. cruziis mediated by membrane pore formation and the induction of nuclear and mitochondrial DNA fragmentation, leading to trypanosome destruction. Exposure of trypomastigote and amastigote forms ofT. cruzito defensin α-1 significantly reduced parasite viability in a peptide concentration-dependent and saturable manner. The toxicity of defensin α-1 againstT. cruziis blocked by anti-defensin α-1 immunoglobulin G. Electron microscopic analysis of trypomastigotes exposed to defensin α-1 revealed pore formation in the cellular and flagellar membranes, membrane disorganization, and blebbing as well as cytoplasmic vacuolization. Furthermore, human defensin α-1 enters the trypanosome when membrane pores are present and is associated with later intracellular damage. Trypanosome membrane depolarization abolished the toxicity of defensin α-1 against the parasite. Preincubation of trypomastigotes with defensin α-1 followed by exposure to human epithelial cells significantly reducedT. cruziinfection in these cells. Thus, human defensin α-1 is an innate immune molecule that causes severe toxicity toT. cruziand plays an important role in reducing cellular infection. This is the first report showing that human defensin α-1 causes membrane pore formation in a human parasite, leading to trypanosome destruction.
Translocation of ExsE into Chinese Hamster Ovary Cells Is Required for Transcriptional Induction of the
Pseudomonas aeruginosa
Type III Secretion System
Authors: Urbanowski Mark L. , Brutinel Evan D. , Yahr Timothy L. 
Published: July 2007

ABSTRACT

Transcription of the
Pseudomonas aeruginosa
type III secretion system (T3SS) is induced under Ca
2+
-limiting growth conditions or following the contact of the bacteria with host cells. The regulatory response to low Ca
2+
levels is initiated by the T3SS-mediated secretion of ExsE, a negative regulatory protein that prevents T3SS gene transcription. In the present study, we demonstrated that ExsE plays an analogous role in transcriptional induction following host cell contact. By using a flow cytometry assay, the host contact-dependent induction of T3SS gene expression was found to be dependent upon the presence of functional type III translocation machinery. Using three independent assays, we demonstrated that ExsE was translocated into Chinese hamster ovary cells in a T3SS-dependent manner. Deletion mapping experiments indicated that the amino terminus of ExsE is required both for secretion under Ca
2+
-limiting growth conditions and for translocation into host cells. A
P. aeruginosa
mutant expressing an
exsE
allele lacking codons 3 through 20 was deficient in ExsE secretion and translocation and showed constitutive repression of T3SS gene expression under Ca
2+
-limiting growth conditions. The mutant also failed to induce T3SS gene expression following host cell contact and demonstrated a significant reduction in T3SS-dependent cytotoxicity towards Chinese hamster ovary cells, indicating that the translocation of ExsE is required for the host contact-dependent induction of T3SS gene expression.
Influence of Host Immunity on Parasite Diversity inTheileria parva
Authors: Katzer Frank , Ngugi Daniel , Schnier Christian , Walker Alan R. , McKeever Declan J. 
Published: July 2007

ABSTRACTWe examined the influence of host immunity on the genotypic diversity of the intracellular transforming cattle parasiteTheileria parva. By tracking the emergence of discrete parasite genotypes in an animal challenged with a bulk stabilate following immunization with its major component clone, we observed a profound modulation of genotypic frequencies in the breakthrough schizont population. In particular, no incidences of the immunizing clone were observed and a progressive decline was apparent in the relatedness of breakthrough genotypes to it. These observations were reflected in the genotypic profile of transmissible parasite stages that emerged in the erythrocyte fraction of the animal and in parasite progeny generated by tick pickup. In a separate experiment, genotypic profiles of breakthrough parasite populations were observed to vary between unrelated immune animals selected on the basis of the major histocompatibility complex (MHC) class I phenotype, a known determinant of the specificity of the immune response. Furthermore, immunization and challenge of calves with molecularly distinct but cross-protective parasite populations revealed that infection results in transmissible erythrocyte forms in spite of a protective immune response. These observations suggest that immunity does not prevent transmission of challenge parasites and that its impact on the parasite at a population level is influenced by herd MHC diversity.
Role for
sagA
and
siaA
in Quorum Sensing and Iron Regulation in
Streptococcus pyogenes
Authors: Salim Kowthar Y. , de Azavedo Joyce C. , Bast Darrin J. , Cvitkovitch Dennis G. 
Published: July 2007

ABSTRACT

Streptococcus pyogenes
is a ubiquitous and versatile pathogen that causes a variety of infections with a wide range of severity. The versatility of this organism is due in part to its capacity to regulate virulence gene expression in response to the many environments that it encounters during an infection. We analyzed the expression of two potential virulence factors,
sagA
and
siaA
(also referred to as
pel
and
htsA
, respectively), in response to conditions of varying cell densities and iron concentrations. The
sagA
gene was up-regulated in conditioned medium from a wild-type strain but not from
sagA
-deficient mutants, and the gene was also up-regulated in the presence of streptolysin S (SLS), the gene product of
sagA
, thus indicating that this gene or its product is involved in density-dependent regulation of
S. pyogenes
. By comparison,
siaA
responded in a manner consistent with a role in iron acquisition since it was up-regulated under iron-restricted conditions. Although
siaA
expression was also up-regulated in the presence of SLS and in conditioned media from both wild-type and
sagA
-deficient mutants, this up-regulation was not growth phase dependent. We conclude that
sagA
encodes a quorum-sensing signaling molecule, likely SLS, and further support the notion that
siaA
is likely involved in iron acquisition.
The E2F-Regulated Gene Chk1 Is Highly Expressed in Triple-Negative Estrogen Receptor−/Progesterone Receptor−/HER-2− Breast Carcinomas
Authors: Verlinden Lieve , Vanden Bempt Isabelle , Eelen Guy , Drijkoningen Maria , Verlinden Ilse , Marchal Kathleen , De Wolf-Peeters Christiane , Christiaens Marie-Rose , Michiels Luc , Bouillon Roger , Verstuyf Annemieke 
Published: July 2007

Abstract
We previously showed that checkpoint kinase 1 (Chk1) and Claspin, two DNA-damage checkpoint proteins, were down-regulated by 1,25-dihydroxyvitamin D3, a known inhibitor of cell proliferation. In the present study, we aimed to investigate the transcriptional regulation of Chk1 and Claspin and to study their expression levels in human breast cancer tissue. Transient transfection experiments in MCF-7 breast cancer cells showed that promoter activities of Chk1 and Claspin were regulated by the E2F family of transcription factors. Subsequently, transcript levels of Chk1, Claspin, and E2F1 were determined by quantitative reverse transcriptase-PCR analysis in 103 primary invasive breast carcinomas and were compared with several clinicopathologic variables in breast cancer. A strong correlation was found between Chk1 and Claspin transcript levels. Transcript levels of Chk1, Claspin, and E2F1 were highest in histologic grade 3 tumors and in tumors in which the expression of estrogen receptor (ER) and progesterone receptor (PR) was lost. Moreover, Chk1 expression was significantly elevated in grade 3 breast carcinomas showing a triple-negative ER−/PR−/HER-2− phenotype compared with other grade 3 tumors. Further research is warranted to validate the use of Chk1 inhibitors in triple-negative breast carcinomas for which treatment strategies are limited at present. [Cancer Res 2007;67(14):6574–81]
The Vascular Disrupting Agent, DMXAA, Directly Activates Dendritic Cells through a MyD88-Independent Mechanism and Generates Antitumor Cytotoxic T Lymphocytes
Authors: Wallace Africa , LaRosa David F. , Kapoor Veena , Sun Jing , Cheng Guanjun , Jassar Arminder , Blouin Aaron , Ching Lai-Ming , Albelda Steven M. 
Published: July 2007

Abstract5,6-Di-methylxanthenone-4-acetic acid (DMXAA) is a small molecule in the flavanoid class that has antitumor activity. Although classified as a “vascular disrupting agent,” we have recently conducted studies showing that DMXAA has remarkable efficacy in a range of tumors, working primarily as an immune modulator that activates tumor-associated macrophages and induces a subsequent CD8+ T-cell–mediated response. To more completely analyze the effect of DMXAA on CD8+ T-cell generation, we treated mice bearing tumors derived from EG7 thymoma cells that express the well-characterized chicken ovalbumin neotumor antigen. Treatment with DMXAA led to cytokine release, tumor cell necrosis, and ultimately reduction in tumor size that was lymphocyte dependent. Within 24 h of administration, we observed dendritic cell activation in tumor-draining lymph nodes (TDLN). This was followed by a rapid and marked increase in the number of tetramer-specific CD8+ T cells in the spleens of treated animals. In contrast, the vascular disrupting agent combretastatin A4-phosphate, which caused a similar amount of immediate tumor necrosis, did not activate dendritic cells, nor induce an effective antitumor response. Using in vitro systems, we made the observation that DMXAA has the ability to directly activate mouse dendritic cells, as measured by increased expression of costimulatory molecules and proinflammatory cytokine release via a pathway that does not require the Toll-like receptor adaptor molecule MyD88. DMXAA thus has the ability to activate tumor-specific CD8+ T cells through multiple pathways that include induction of tumor cell death, release of stimulatory cytokines, and direct activation of dendritic cells. [Cancer Res 2007;67(14):7011–9]
Subcellular Localization of Sigma-2 Receptors in Breast Cancer Cells Using Two-Photon and Confocal Microscopy
Authors: Zeng Chenbo , Vangveravong Suwanna , Xu Jinbin , Chang Katherine C. , Hotchkiss Richard S. , Wheeler Kenneth T. , Shen Duanwen , Zhuang Zhi-Ping , Kung Hank F. , Mach Robert H. 
Published: July 2007

AbstractSigma-2 receptor agonists have been shown to induce cell death via caspase-dependent and caspase-independent pathways. Unfortunately, there is little information regarding the molecular function of sigma-2 receptors that can explain these results. In this study, two fluorescent probes, SW107 and K05-138, were used to study the subcellular localization of sigma-2 receptors by two-photon and confocal microscopy. The results indicate that sigma-2 receptors colocalize with fluorescent markers of mitochondria, lysosomes, endoplasmic reticulum, and the plasma membrane in both EMT-6 mouse and MDA-MB-435 human breast cancer cells. The fluorescent probe, K05-138, was internalized rapidly, reaching a plateau of fluorescent intensity at 5 min. The internalization of K05-138 was reduced ∼40% by phenylarsine oxide, an inhibitor of endocytosis. These data suggest that sigma-2 ligands are internalized, in part, by an endocytotic pathway. The localization of sigma-2 receptors in several organelles known to have a role in both caspase-dependent and caspase-independent pathways of cell death supports the conclusions of previous studies suggesting that sigma-2 receptor ligands should be evaluated as potential cancer chemotherapeutic agents. [Cancer Res 2007;67(14):6708–16]
Tumor Paint: A Chlorotoxin:Cy5.5 Bioconjugate for Intraoperative Visualization of Cancer Foci
Authors: Veiseh Mandana , Gabikian Patrik , Bahrami S-Bahram , Veiseh Omid , Zhang Miqin , Hackman Robert C. , Ravanpay Ali C. , Stroud Mark R. , Kusuma Yumiko , Hansen Stacey J. , Kwok Deborah , Munoz Nina M. , Sze Raymond W. , Grady William M. , Greenberg Norman M. , Ellenbogen Richard G. , Olson James M. 
Published: July 2007

Abstract
Toward the goal of developing an optical imaging contrast agent that will enable surgeons to intraoperatively distinguish cancer foci from adjacent normal tissue, we developed a chlorotoxin:Cy5.5 (CTX:Cy5.5) bioconjugate that emits near-IR fluorescent signal. The probe delineates malignant glioma, medulloblastoma, prostate cancer, intestinal cancer, and sarcoma from adjacent non-neoplastic tissue in mouse models. Metastatic cancer foci as small as a few hundred cells were detected in lymph channels. Specific binding to cancer cells is facilitated by matrix metalloproteinase-2 (MMP-2) as evidenced by reduction of CTX:Cy5.5 binding in vitro and in vivo by a pharmacologic blocker of MMP-2 and induction of CTX:Cy5.5 binding in MCF-7 cells following transfection with a plasmid encoding MMP-2. Mouse studies revealed that CTX:Cy5.5 has favorable biodistribution and toxicity profiles. These studies show that CTX:Cy5.5 has the potential to fundamentally improve intraoperative detection and resection of malignancies. [Cancer Res 2007;67(14):6882–8]
Effect of Dominant Negative Transforming Growth Factor-β Receptor Type II on Cytotoxic Activity of RAW 264.7, a Murine Macrophage Cell Line
Authors: Lee Geun Taek , Hong Jun Hyuk , Kwak Cheol , Woo Jaesung , Liu Victoria , Lee Chung , Kim Isaac Yi 
Published: July 2007

AbstractTransforming growth factor-β (TGF-β) is a potent suppressor of the immune system. In the present study, we investigated the effect of TGF-β resistance on a murine macrophage cell line, RAW 264.7, by overexpressing a dominant negative TGF-β receptor type II (TβRIIDN) construct. As expected, TβRIIDN-expressing RAW cells, designated as RAW-TβRIIDN, were resistant to TGF-β signaling. When these cells were cocultured with the murine renal cell carcinoma cell line, Renca, a dramatic increase in apoptosis of Renca cells was observed. Simultaneously, elevated levels of inducible nitric oxide synthase (iNOS) and tumor necrosis factor-α (TNF-α) in association with IFN-γ were detected in RAW-TβRIIDN cells. When the effects of TNF-α and iNOS were neutralized through the use of neutralizing antibody and NG-methyl-l-arginine, respectively, the enhanced cytotoxicity of TβRIIDN-RAW cells was partially reversed. Taken together, these results show that TGF-β–resistant RAW 264.7 murine macrophage cells have increased cytotoxic activity that is in part mediated by iNOS and TNF-α. [Cancer Res 2007;67(14):6717–24]
Inhibition of Nuclear Factor-κB Activity by Temozolomide Involves O6-Methylguanine–Induced Inhibition of p65 DNA Binding
Authors: Yamini Bakhtiar , Yu Xiaohong , Dolan M. Eileen , Wu Michael H. , Kufe Donald W. , Weichselbaum Ralph R. 
Published: July 2007

Abstract
The alkylating agent temozolomide, commonly used in the treatment of malignant glioma, causes cellular cytotoxicity by forming O6-methylguanine adducts. In this report, we investigated whether temozolomide alters the activity of the transcription factor nuclear factor-κB (NF-κB). Temozolomide inhibits basal and tumor necrosis factor α (TNFα)–induced NF-κB transcriptional activity without altering phosphorylation or degradation of inhibitor of κB-α. Inhibition of NF-κB is secondary to attenuation of p65 DNA binding, not nuclear translocation. Inhibition of DNA binding is shown both in vitro, with gel shift studies and DNA binding assays, and in vivo at κB sites. Consistent with inhibition of NF-κB activity, temozolomide reduces basal and TNFα-induced κB-dependent gene expression. Temozolomide also inhibits NF-κB activated by inducers other than TNFα, including lipopolysaccharide, doxorubicin, and phorbol 12-myristate 13-acetate. The inhibitory action of temozolomide on NF-κB is observed to be maximal following pretreatment of cells with temozolomide for 16 h and is also seen with the SN1-type methylating agent methylnitrosourea. The ability of temozolomide to form O6-methylguanine adducts is important for inhibition of NF-κB as is the presence of a functioning mismatch repair system. Activation of NF-κB with TNFα before administration of temozolomide reduces the cytotoxicity of temozolomide, whereas 16-h pretreatment with temozolomide resensitizes cells to killing. This work shows a mechanism whereby O6-methylguanine adducts formed by temozolomide lead to inhibition of NF-κB activity and illustrates a link between mismatch repair processing of alkylator-induced DNA damage and cell death. [Cancer Res 2007;67(14):6889–98]