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Director: Sunita Seemanapalli (sseema1@lsu.edu)

Focus: Bacteriology, Molecular biology, Immunology

Description:

Schedules:

2009-09-18 14:00:00 UTC: Temperature-induced regulation of RpoS by a small RNA in Borrelia burgdorferi - organized by Yihe Ge

Temperature-induced regulation of RpoS by a small RNA in Borrelia burgdorferi Disgussion

Summary: The alternative sigma factor RpoS (sigma38 or sigmaS) plays a central role in the reciprocal regulation of the virulence-associated major outer surface proteins OspC and OspA in Borrelia burgdorferi, the Lyme disease spirochete. Temperature is one of the key environmental signals controlling RpoS, but the molecular mechanism by which the signal is transduced remains unknown. Herein, we identify and describe a small non-coding RNA, DsrABb, that regulates the temperature-induced increase in RpoS. A novel 5' end of the rpoS mRNA was identified and DsrABb has the potential to extensively base-pair with the upstream region of this rpoS transcript. We demonstrate that B. burgdorferi strains lacking DsrABb do not upregulate RpoS and OspC in response to an increase in temperature, but do regulate RpoS and OspC in response to changes in pH and cell density. Analyses of the rpoS and ospC steady-state mRNA levels in the dsrABb mutant indicate that DsrABb regulates RpoS post-transcriptionally. The 5' and 3' ends of DsrABb were mapped, demonstrating that at least four species exist with sizes ranging from 213 to 352 nucleotides. We hypothesize that DsrABb binds to the upstream region of the rpoS mRNA and stimulates translation by releasing the Shine-Dalgarno sequence and start site from a stable secondary structure. Therefore, we postulate that DsrABb is a molecular thermometer regulating RpoS in Borrelia burgdorferi.

2009-08-20 09:00:00 UTC: OspC-Independent Infection and Dissemination by Host-Adapted Borrelia burgdorferi - organized by poonam dadhwal

OspC-Independent Infection and Dissemination by Host-Adapted Borrelia burgdorferi Disgussion

Summary: Borrelia burgdorferi OspC is required for the spirochete to establish infection in a mammal by tick transmission or needle inoculation. After a brief essential period, the protein no longer is required and the gene can be shut off. Using a system in which spirochetes contain only an unstable wild-type copy of the ospC gene, we can obtain mice persistently infected with bacteria lacking OspC. We implanted pieces of infected mouse skin subcutaneously in naïve mice, using donors carrying wild-type or ospC mutant spirochetes, and found that both could infect mice by this method, with similar numbers of wild-type or ospC mutant spirochetes disseminated throughout the tissues of recipient mice. Recipient mouse immune responses to tissue transfer-mediated infection with wild-type or ospC mutant spirochetes were similar. These experiments demonstrate that mammalian host-adapted spirochetes can infect and disseminate in mice in the absence of OspC, thereby circumventing this hallmark of tick-derived or in vitro-grown spirochetes. We propose a model in which OspC is one of a succession of functionally equivalent, essential proteins that are synthesized at different stages of mammalian infection. In this model, another protein uniquely present on host-adapted spirochetes performs the same essential function initially fulfilled by OspC. The strict temporal control of B. burgdorferi outer surface protein gene expression may reflect immunological constraints rather than distinct functions.

2009-08-13 09:00:00 UTC: Application of chicken microarrays for gene expression analysis in other avian species - organized by Rahul Sharma

Application of chicken microarrays for gene expression analysis in other avian species Disgussion

Summary: With the threat of emerging infectious diseases such as avian influenza, whose natural hosts are thought to be a variety of wild water birds including duck, we are armed with very few genomic resources to investigate large scale immunological gene expression studies in avian species. Multiple options exist for conducting large gene expression studies in chickens and in this study we explore the feasibility of using one of these tools to investigate gene expression in other avian species. In this study we utilised a whole genome long oligonucleotide chicken microarray to assess the utility of cross species hybridisation (CSH). We successfully hybridised a number of different avian species to this array, obtaining reliable signals. We were able to distinguish ducks that were infected with avian influenza from uninfected ducks using this microarray platform. In addition, we were able to detect known chicken immunological genes in all of the hybridised avian species. Cross species hybridisation using long oligonucleotide microarrays is a powerful tool to study the immune response in avian species with little available genomic information. The present study validated the use of the whole genome long oligonucleotide chicken microarray to investigate gene expression in a range of avian species.

2009-08-06 09:00:00 UTC: NOD1 activates CEBP beta signaling in Lung epithelial cells - organized by Kohila Mahadevan

The pattern recognition receptor Nod1 activates CCAAT/enhancer binding protein b signalling in lung epithelial cells Disgussion

Summary: The innate immune receptor nucleotide-binding oligomerisation domain protein 1 (Nod1) recognises peptidoglycan containing meso-diaminopimelic acid found in all Gramnegative and some Gram-positive bacteria. Nod1 has been shown to activate nuclear factor (NF)- kB. The aim of the present study was to examine the expression of Nod1 in the lung, particularly in lung epithelial cells, and to investigate the activation of CCAAT/enhancer binding protein (C/EBP) transcription factors downstream of the Nod1 receptor in these cells. The expression of Nod1 in mouse lung was examined using immunohistochemistry. A tissue array was used to determine the expression pattern in the human lung. Signalling downstream of Nod1 was examined in the human lung epithelial cell type, BEAS-2B, by electrophoretic mobility shift assay and reporter gene activation. Nod1 expression was seen in various cell types in the lung, including epithelial cells. Activation of Nod1 in these cells resulted in modest activation of NF-kB, together with strong activation of the C/EBP transcription factors, particularly C/EBPb. This activation appears to be independent of de novo protein synthesis. The present study showed that nucleotide-binding oligomerisation domain protein 1 is expressed in lung epithelial cells. The results demonstrate a novel pathway downstream of the nucleotide-binding oligomerisation domain protein 1 receptor in these cells and suggest that C/ EBPb may play a role in immune responses to meso-diaminopimelic acid-containing bacteria in the lung.

2009-07-16 09:00:00 UTC: Lung NF-B Activation and Neutrophil Recruitment Require - organized by Gayathriy Balamayooran

Lung NF-B Activation and Neutrophil Recruitment Require IL-1 and TNF Receptor Signaling during Pneumococcal Pneumonia Disgussion

Summary: Pulmonary inflammation is an essential component of the host defense against Streptococcus pneumoniae infection of the lungs. The early response cytokines, TNF- and IL-1, are rapidly induced upon microbial exposure. Mice deficient in all TNF- and IL-1-dependent signaling receptors were used to determine the roles of these cytokines during pneumococcal pneumonia. The deficiency of signaling receptors for TNF and IL-1 decreased bacterial clearance. Neutrophil recruitment to alveolar air spaces was impaired by receptor deficiency, as was pulmonary expression of the neutrophil chemokines KC and MIP-2. Because NF-B mediates the expression of both chemokines, we assessed NF-B activation in the lungs. During pneumococcal pneumonia, NF-B proteins translocate to the nucleus and activate gene expression; these functions were largely abrogated by the deficiency of receptors for TNF- and IL-1. Thus, the combined deficiency of TNF and IL-1 signaling reduces innate immune responses to S. pneumoniae in the lungs, probably due to essential roles for these receptors in activating NF-B.

2009-07-01 14:49:00 UTC: the differential expression of subolesin in ticks in response to anaplasma infection - organized by Piyanate Sunyakumthorn

the differential expression of tick genes in response to anaplasma infection. Disgussion

Summary: Subolesin is a protective antigen that was discovered in ticks. The subolesin gene is conserved in invertebrates and vertebrates and is found in many tick genera. Recently, the study of RNA interference and immunization show the protection of tick infestations by reduction of tick survival, feeding, and reproduction. Moreover, the blocking of subolesin by RNAi and vaccination resulted in a decrease of Anaplasma infection in the host cells. The results from this article (De la fuente, 2008) support the role of subolesin in an Anaplasma infected tick and suggest that subolesin can be a good candidate for vaccine antigen against tick infestation and also Anaplasma infection.

2009-06-25 01:18:00 UTC: Chemokine and cytokine processing by MMP - organized by Gayathriy Balamayooran

Chemokine and cytokine processing by matrix Disgussion

Summary: The action of matrix metalloproteinases (MMPs) was originally believed to be restricted to degradation of the extracellular matrix; however, in recent years, it has become evident that these proteases can modify many nonmatrix substrates, such as cytokines and chemokines. The use of MMP-deficient animals has revealed that these proteases can indeed influence the progression of various inflammatory processes. This review aims to provide the reader with a concise overview of these novel MMP functions in relation to leukocyte migration.

2009-06-18 09:00:00 UTC: 2-component regulatory system - Treponema denticola - organized by Sunita Seemanapalli

Analysis of a Growth-Phase-Regulated Two-Component Regulatory System in the Periodontal Pathogen Treponema denticola Disgussion

Summary: Nothing is currently known regarding the global regulatory networks of Treponema denticola and other oral spirochetes. In this report, we assess the properties and potential phosphotransfer capability of a putative two-component regulatory system (TCS) of T. denticola that is formed by the products of open reading frames tde0032 (a sensor kinase) and tde0033 (a response regulator), henceforth designated AtcS and AtcR, respectively. Using PCR and DNA sequence analyses, atcS and atcR were demonstrated to be widely distributed and conserved among T. denticola isolates. Reverse transcription-PCR (RT-PCR) analyses revealed that these genes are cotranscribed and may also be expressed as part of a larger operon that includes several flanking genes. Analyses using 5 rapid amplification of cDNA ends identified the transcriptional start sites for these operons and provided evidence that some of these genes may be independently transcribed from internal promoters. Real-time RT-PCR and Western blot analysis revealed significant upregulation of atcRS during late-stage growth, indicating growth-phase-dependent expression. Lastly, the phosphorelay capability of the AtcRS system was assessed and demonstrated using recombinant proteins. AtcS was found to undergo autophosphorylation and to transfer phosphate to AtcR. These analyses represent the first description of a functional TCS in an oral spirochetes and provide insight into the transcriptional regulatory mechanisms of these important bacteria.

2009-06-05 17:30:00 UTC: Inflammasome - organized by Kohila Mahadevan

2009-06-04 14:11:00 UTC: This is the another test - organized by Qilong Xu

2009-06-04 09:00:00 UTC: Inflammasome - Test - organized by Sunita Seemanapalli

2009-04-03 09:00:00 UTC: Innovative Technologies - organized by Sunita Seemanapalli

Two-in-one designer antibodies Disgussion

Summary: An antibody is engineered to recognize two different proteins with high affinity, opening the door to improved combination therapies for cancers and infections

2009-03-20 09:00:00 UTC: Mucosal Immunity - organized by Gayathriy Balamayooran

Mucosal immunity and vaccines Disgussion

Summary: Deals with the recent interest in vaccine development to promote mucosal immunity

2009-03-06 05:19:00 UTC: Toxoid Vaccines and Passive immunity - organized by Gayathriy Balamayooran

Basics of toxoid vaccines and importance of Passive immunity Disgussion

Summary: Toxoid vaccines are mainly used in bacterial diseases in which severe pathology is caused by exotoxins released by the bacteria. Tetanus, diptheria and pertusis are such important diseases. Toxoid vaccine are made of inactivated exotoxin.......................................... Passive immunity is the transfer of antibodies from an immuned organism to and non immuned organism. This can occur naturally as well as artificially. This immunity is of importance when the organism is immunologically immature or in as immunosuppressed state.

2009-02-20 09:00:00 UTC: DNA Vaccines - organized by Qilong Xu

DNA Vaccines-overview Disgussion

Summary: The DNA vaccines are simple rings of DNA containing a gene encoding an antigen, and a promoter/terminator to make the gene express in mammalian cells. We will discuss the principle, mechanisms, limitations, and perspective of DNA vaccines.

2009-02-06 09:00:00 UTC: Lab work and projects - organized by Sunita Seemanapalli

2009-01-29 09:00:00 UTC: Vaccines continues... - organized by Kohila Mahadevan

2009-01-23 09:00:00 UTC: Vaccines-In general - organized by Sunita Seemanapalli

Vaccines against intracellular bacterial pathogens Richard W. Titball School of Biosciences, University of Exeter, Exeter, EX4 4QD Devon, UK Disgussion

Summary: There is a long history of remarkable success in developing vaccines against bacteria that are extracellular pathogens. In general, the development of vaccines against intracellular bacterial pathogens has proven to be more challenging. Typically, such vaccines need to induce a range of immune responses, including antibody, CD4+ and CD8+ T cell responses. These responses can be induced by live attenuated vaccines, but eliciting these responses with non-living vaccines has proven to be difficult. The difficulties appear to be related partly to the problems associated with the identification of protective antigens and partly with the difficulties associated with inducing CD8+ T cell responses.

Vaccines, Vaccination, and Vaccinology Stanley A. Plotkin Department of Pediatrics, University of Pennsylvania, Philadelphia, and Aventis Pasteur, Doylestown, Pennsylvania The Journal of Infectious Diseases 2003;187:1349–59 Disgussion

Summary: Although the demonstration in 1796 by Edward Jenner that vaccinia virus could protect against smallpox was epochal, he was following the path opened by the ancients who had used the smallpox virus itself in the practice of variolation. The work of Louis Pasteur on chicken cholera opened the way to vaccine development in the laboratory. In April 1880, Pasteur reported to the French Academy of Science that “…chicken cholera is produced by a microscopic parasite [now known as Pasteurella multocida], that there exists an attenuated virus [Pasteur was using “virus” in the ancient sense of the word] of that disease, and that one or more inoculations of this attenuated virus can preserve the animals from the mortal effects of a later inoculation … let me be permitted to use the word ‘vaccinate’ to express the act of inoculating a chicken with the attenuated virus” [1]. Thus, the word “vaccinate” was extended beyond vaccinia and came to have its modern meaning. Today, epidemic infectious diseases of children for which there are vaccines have virtually disappeared from industrialized countries such as the United States. These are remarkable successes, which together with clean water and antibiotics have profoundly affected human society [2]. In addition, a new field of microbiology and immunology has evolved, called “vaccinology,” that comprises not only vaccine development but also the use of vaccines and their effects on public health [3]. Here, I will briefly cover some aspects of the past, discuss 5 current issues in vaccinology,and then turn to the future.

Pathogens-Vaccines Disgussion

Summary: Pathogens: Intracellular, Facultative Intracellular Pathogens, Extracellular Antigen presentation Vaccines against intracellular and extracellular pathogens

2008-12-05 09:00:00 UTC: RNA-binding regulatory proteins - organized by Qilong Xu

CsrB sRNA family sequestration of RNA-binding regulatory Disgussion

Summary: Noncoding regulatory RNA molecules, also known as small RNAs, participate in several bacterial regulatory networks. The central component of the carbon storage regulator (Csr) and the homologous repressor of secondary metabolites (Rsm) systems is an RNA binding protein (CsrA or RsmA) that regulates gene expression post-transcriptionally by affecting ribosome binding and/or mRNA stability. Members of the CsrB family of noncoding regulatory RNA molecules contain multiple CsrA binding sites and function as CsrA antagonists by sequestering this protein. Depending on the particular organism, the Csr (or Rsm) system participates in global regulatory circuits that control central carbon ﬂux, the production of extracellular products, cell motility, bioﬁlm formation, quorum sensing and/or pathogenesis.

Gac-Rsm signal transduction pathway of proteobacteria Disgussion

Summary: In many g-proteobacteria, the conserved GacS/GacA (BarA/UvrY) two-component system positively con- trols the expression of one to ﬁve genes specifying small RNAs (sRNAs) that are characterized by repeated unpaired GGA motifs but otherwise appear to belong to several independent families. The GGA motifs are essential for binding small, dimeric RNA- binding proteins of a single conserved family desig- nated RsmA (CsrA). These proteins, which also occur in bacterial species outside the g-proteobacteria, act as translational repressors of certain mRNAs when these contain an RsmA/CsrA binding site at or near the Shine-Dalgarno sequence plus additional binding sites located in the 5' untranslated leader mRNA. Recent structural data have established that the RsmA-like protein RsmE of Pseudomonas ﬂuore- scens makes speciﬁc contacts with an RNA consen- sus sequence 5'-A/UCANGGANGU/A-3' (where N is any nucleotide). Interaction with an RsmA/CsrA protein promotes the formation of a short stem supporting an ANGGAN loop. This conformation hinders access of 30S ribosomal subunits and hence translation initiation. The output of the Gac/Rsm cascade varies widely in different bacterial species and typically involves management of carbon storage and expres- sion of virulence or biocontrol factors. Unidentiﬁed signal molecules co-ordinate the activity of the Gac/ Rsm cascade in a cell population density-dependent manner.

Bacterial Small RNA Regulators Disgussion

Summary: All about small RNA regulators.

2008-11-19 09:00:00 UTC: Toll-like receptors-module I - organized by Qilong Xu

Distinct roles for MyD88, TLR2, TLR5, and TLR9 in phagocytosis of... Disgussion

Summary: The contribution of toll-like receptors (TLRs) to phagocytosis of Borrelia burgdorferi has not been extensively studied. We show that bone marrow derived macrophages (BMDM) from MyD88-/- mice or Raw cells transfected with a dominant negative MyD88 were unable to efficiently internalize B. burgdorferi. Knockouts of TLR2 and TLR9 or knockdown of TLR5 by siRNA produced no defects in phagocytosis of B. burgdorferi. Production of inflammatory cytokines was greatly diminished in MyD88-/- BMDM, but only partially affected in TLR2-/- BMDM or knockdown of TLR5 and unaffected in TLR9-/- BMDM. Cytochalasin D (cytoD) reduced cytokine induction, but not to the level of the MyD88-/-. Addition of cytoD to TLR2-/- BMDM inhibited inflammatory responses to B. burgdorferi to the level of MyD88-/- BMDM-- consistent with a role for TLR2 in both recognition of extracellular products and lysosomal sampling by TLR2 after processing of the organism. CytoD had no impact on cytokine productions in cells undergoing TLR5 knockdown. These results suggest that MyD88, but not TLR2, TLR5 and TLR9, is important for the uptake of B. burgdorferi and MyD88 affects inflammatory responses both through its effects on phagocytosis and its role in transducing signals from TLR2 and TLR5.

TLR signaling pathways Disgussion

Summary: Toll-like receptors (TLRs) have been established to play an essential role in the activation of innate immunity by recognizing spe- ciﬁc patterns of microbial components. TLR signaling pathways arise from intracytoplasmic TIR domains, which are conserved among all TLRs. Recent accumulating evidence has demonstrated that TIR domain-containing adaptors, such as MyD88, TIRAP, and TRIF, modulate TLR signaling pathways. MyD88 is essential for the induction of inﬂammatory cytokines triggered by all TLRs. TIRAP is speciﬁcally involved in the MyD88-dependent pathway via TLR2 and TLR4, whereas TRIF is implicated in the TLR3- and TLR4-mediated MyD88-independent pathway. Thus, TIR domain-containing adaptors provide speciﬁcity of TLR signaling.

2008-11-14 09:00:00 UTC: Immunology-Review - organized by Sunita Seemanapalli

Immunoglobulins Disgussion

Summary: immunoglobulins generally assume one of two roles: immunoglobulins may act as i) plasma membrane bound antigen receptors on the surface of a B-cell or ii) as antibodies free in cellular fluids functioning to intercept and eliminate antigenic determinants. In either role, antibody function is intimately related to its structure and this page will introduce immunoglobulins (antibodies) and relate their structure to their function in host defense.

2008-11-07 09:00:00 UTC: Adaptive Immunity - organized by Sunita Seemanapalli

Activation of CD4 T cells Disgussion

Summary: Physical detection of antigen-speciﬁc CD4 T cells has revealed fea- tures of the in vivo immune response that were not appreciated from in vitro studies. In vivo, antigen is initially presented to na¨ıve CD4 T cells exclusively by dendritic cells within the T cell areas of secondary lymphoid tissues. Anatomic constraints make it likely that these dendritic cells acquire the antigen at the site where it en- ters the body. Inﬂammation enhances in vivo T cell activation by stimulating den- dritic cells to migrate to the T cell areas and display stable peptide-MHC complexes and costimulatory ligands. Once stimulated by a dendritic cell, antigen-speciﬁc CD4 T cells produce IL-2 but proliferate in an IL-2–independent fashion. Inﬂammatory signals induce chemokine receptors on activated T cells that direct their migration into the B cell areas to interact with antigen-speciﬁc B cells. Most of the activated T cells then die within the lymphoid tissues. However, in the presence of inﬂam- mation, a population of memory T cells survives. This population is composed of two functional classes. One recirculates through nonlymphoid tissues and is capable of immediate effector lymphokine production. The other recirculates through lymph nodes and quickly acquires the capacity to produce effector lymphokines if stimulated. Therefore, antigenic stimulation in the presence of inﬂammation produces an increased number of speciﬁc T cells capable of producing effector lymphokines throughout the body.

Borrelia burgdorferi–Infected, Interleukin-6–Deﬁcient Mice Disgussion

Summary: Recently, interleukin (IL)-6 was shown to be one of the earliest factors that trigger the differentiation of naive T cells into effector Th2 cells in vitro. Lyme arthritis was studied in IL-6–deﬁcient mice, since joint inﬂammation is inﬂuenced by the T helper cell response against Borrelia burgdorferi. Arthritis incidence increased in B. burgdorferi–infected IL-6– deﬁcient mice compared with that in controls. Furthermore, splenocytes of B. burgdor- feri–infected IL-6–deﬁcient mice produced signiﬁcantly less IL-4 in response to Borrelia an- tigens than did C57BL/6 (B6) mice, and B. burgdorferi–speciﬁc IgG2b levels were signiﬁcantly reduced in IL-6–deﬁcient mice at 60 days of infection. These results extend previous in vitro observations by demonstrating an in vivo role for IL-6 in the differentiation of CD4 T cells toward a Th2 phenotype and further show that CD4 T cell responses inﬂuence murine Lyme arthritis.

Club members: Send an invitation

Sunita Seemanapalli : sseema1@lsu.edu

Qilong Xu : qxu@vetmed.lsu.edu

Yihe Ge : geyihe@lsu.edu

GAYATHRIY BALAMAYOORAN : bgay.maha@gmail.com

Suraj narayan Shrestha : surajnstha@hotmail.com

Balamayooran Theivanthiran : balamayooran5@gmail.com

Kohila Mahadevan : kmahad1@tigers.lsu.edu

Gayathriy Balamayooran : gmahad1@tigers.lsu.edu.com

Piyanate Sunyakumthorn : psunyaku@tigers.lsu.edu

balamayooran theivanthiran : mayoorant@yahoo.com

poonam dadhwal : pdadhw1@tigers.lsu.edu

sona chowdhury : schowd1@lsu.edu

Rahul Sharma : rsharm4@lsu.edu

xiaomei Hu : xhu@lsu.edu