Involvement of SPI-2-encoded SpiC in flagellum synthesis in Salmonella enterica serovar Typhimurium

Background  

SpiC encoded within Salmonella pathogeniCity island 2 on the Salmonella enterica serovar Typhimurium chromosome is required for survival within macrophages and systemic infection in mice. Additionally, SpiC contributes to Salmonella-induced activation of the signal transduction pathways in macrophages by affecting the expression of FliC, a component of flagella filaments. Here, we show the contribution of SpiC in flagellum synthesis.     

Inhibition of HIV‐1 infection by a unique short hairpin RNA to chemokine receptor 5 delivered into macrophages through hematopoietic progenitor cell transduction

Background

We recently expressed a potent and noncytotoxic short hairpin (sh)RNA directed against chemokine (c‐c motif) receptor 5 (CCR5) using lentiviral mediated transduction of CD34+ hematopoietic progenitor cells (HPCs) and demonstrated the stable reduction of CCR5 expression in T‐lymphocytes.

Methods

In the present study, we further assessed the activity of the shRNA through HPC transduction and differentiation into macrophages derived from fetal liver CD34+ (FL‐CD34+) HPCs. Transduced lentiviral vector encoding the human CCR5 shRNA was stably maintained in FL‐CD34+ cells and in the terminally differentiated macrophages using macrophage colony‐stimulating factor, granulocyte macrophage colony‐stimulating factor, interleukin‐3 and stem cell factor.

Results

Quantitative real‐time polymerase chain reaction for CCR5 mRNA indicated over 90% reduction of CCR5 mRNA levels in CCR5 shRNA‐transduced population. The cells with knockdown of CCR5 expression acquired resistance to R5 tropic HIV‐1 NFN‐SX strain. We also developed a novel approach utilizing a mCherry‐CCR5 chimeric reporter to assess the effectiveness of CCR5 target down‐regulation in macrophages directly. Both the shRNA and the reporter were maintained throughout HPC differentiation to macrophages without apparent cytotoxicity.

Conclusions

The present study demonstrates a novel method to simply and directly assess the function of small interfering RNA and the effective inhibition of HIV‐1 infection by a potential potent shRNA to CCR5 delivered into macrophages derived from HPCs. Copyright © 2010 John Wiley & Sons, Ltd.     

Detecting and minimizing zinc contamination in physiological solutions

Background  

Rat liver endosomes contain activated insulin receptors and downstream signal transduction molecules. We undertook these studies to determine whether endosomes also contain heterotrimeric G proteins that may be involved in signal transduction from G protein-coupled receptors.     

A topological model of biofeedback based on catecholamine interactions

Background  

The present paper describes a topological model of biofeedback. This model incorporates input from a sensory organ and a transduction phase mediated through catecholamine production in the feedback path. The transduction phase comprises both conservative and dissipative systems, from which the appropriate output is combined in a closed loop.     

High efficiency lentiviral gene delivery in non-dividing cells by deoxynucleoside treatment

Background

Gene therapy has recently been advanced by the development of HIV‐based vectors that are able to transduce some non‐dividing cells. The manipulation of most non‐dividing cells remains, however, scarcely efficient. One of the biological mechanisms postulated to prevent powerful transduction of quiescent cells by lentiviral vectors is the paucity of deoxynucleotides (dNTPs). In this study, a novel delivery strategy is developed to improve significantly the efficiency of HIV‐based vectors in transducing non‐dividing cells. This approach is based on increasing the intracellular availability of dNTPs by incubating target cells with the dNTP precursors, deoxynucleosides (dNSs).

Methods

Mature human monocyte‐derived macrophages (14–21 days old) were transduced at a low multiplicity of infection (MOI) of HIV vectors carrying a reporter gene. dNSs were added to the medium during transduction (5 mM dNS) and immediately before post‐transduction culture (2.5 mM dNS). Macrophages were harvested 2–7 days after transduction and assayed for transgene expression by cytofluorimetry.

Results

The addition of dNS to the medium significantly enhanced the efficiency of transduction of human macrophages by HIV‐based vectors. The percentage of cells expressing the transgene rose up to 50% in the presence of dNS, increasing the basal transduction levels up to 35‐fold (average=10.8‐fold). Furthermore, treatment with dNTP precursors compensated for the wide inter‐donor variability, allowing the highest enhancement effects in donors with the lowest basal transduction efficiencies.

Conclusions

This is the first demonstration that a single treatment of non‐dividing target cells with exogenous dNS can enhance the efficiency of lentiviral‐mediated transduction of cells, allowing for high efficiency gene transfer. The effects of dNTP precursors compensated for both the poor basal levels and the wide inter‐donor variability, two major limitations for the transduction of non‐dividing cells. Macrophages are a representative model of cells whose permissiveness to gene delivery was increased up to levels suitable for genetic manipulation applications. This simple approach might be transferred to a broader range of quiescent cell types that are scarcely susceptible to lentiviral‐based gene delivery due to low dNTP levels. Copyright © 2002 John Wiley & Sons, Ltd.

     

Killing of Mycobacterium avium subspecies paratuberculosis within macrophages

Background  

Mycobacterium avium subspecies paratuberculosis (M. paratuberculosis) is a facultative intracellular pathogen that resides within host macrophages during infection of ruminant animals. We examined survival of M. paratuberculosis infections within cultured macrophages to better understand the interplay between bacterium and host.     

Gli2a protein localization reveals a role for Iguana/DZIP1 in primary ciliogenesis and a dependence of Hedgehog signal transduction on primary cilia in the zebrafish

Background  

In mammalian cells, the integrity of the primary cilium is critical for proper regulation of the Hedgehog (Hh) signal transduction pathway. Whether or not this dependence on the primary cilium is a universal feature of vertebrate Hedgehog signalling has remained contentious due, in part, to the apparent divergence of the intracellular transduction pathway between mammals and teleost fish.     

Identification of two Mycobacterium tuberculosis H37Rv ORFs involved in resistance to killing by human macrophages

Background  

The ability of Mycobacterium tuberculosis to survive and replicate in macrophages is crucial for the mycobacterium's ability to infect the host and cause tuberculosis. To identify Mycobacterium tuberculosis genes involved in survival in macrophages, a library of non-pathogenic Mycobacterium smegmatis bacteria, each carrying an individual integrated cosmid containing M. tuberculosis H37Rv genomic DNA, was passed through THP-1 human macrophages three times.     

PathFinder: mining signal transduction pathway segments from protein-protein interaction networks

Background  

A Signal transduction pathway is the chain of processes by which a cell converts an extracellular signal into a response. In most unicellular organisms, the number of signal transduction pathways influences the number of ways the cell can react and respond to the environment. Discovering signal transduction pathways is an arduous problem, even with the use of systematic genomic, proteomic and metabolomic technologies. These techniques lead to an enormous amount of data and how to interpret and process this data becomes a challenging computational problem.     

Chromosomal loci important for cotyledon opening under UV-B in Arabidopsis thaliana

Background  

Understanding of the genetic architecture of plant UV-B responses allows extensive targeted testing of candidate genes or regions, along with combinations of those genes, for placement in metabolic or signal transduction pathways.     

Towards the systematic discovery of signal transduction networks using phosphorylation dynamics data

Background  

Phosphorylation is a ubiquitous and fundamental regulatory mechanism that controls signal transduction in living cells. The number of identified phosphoproteins and their phosphosites is rapidly increasing as a result of recent mass spectrometry-based approaches.     

Apoptosis of resident and inflammatory macrophages before and during the inflammatory response of the virgin bovine mammary gland

Background  

Macrophages may play a prominent role in defense of the bovine mammary gland, and their functionality is necessary for successful eradication of bacterial pathogens. In contrast to necrosis, however, apoptosis has not yet been studied in macrophages from bovine mammary glands. Therefore, the aim of this study was to confirm the occurrence of apoptosis in macrophages from resting heifer mammary glands and during the inflammatory response.     

Transduction of the MPG-tagged fusion protein into mammalian cells and oocytes depends on amiloride-sensitive endocytic pathway

Background  

MPG is a cell-permeable peptide with proven efficiency to deliver macromolecular cargoes into cells. In this work, we examined the efficacy of MPG as an N-terminal tag in a fusion protein to deliver a protein cargo and its mechanism of transduction.     

Function of nuclear transport factor 2 and Ran in the 20E signal transduction pathway in the cotton bollworm, Helicoverpa armigera

Background  

Nuclear transport factor 2 and small GTPase Ran participate in the nucleo-cytoplasm transport of macromolecules, but their function in the 20-hydroxyecdysone (20E) signal transduction pathway are not well known.     

Functional and structural characteristics of anticancer peptide Pep27 analogues

Background  

A secreted peptide Pep27 initiates the cell death program in S. pneumoniae through signal transduction. This study was undertaken to evaluate the relation between the structure and cytotoxic activity of Pep27 and its analogues on cancer cells.     

A census of membrane-bound and intracellular signal transduction proteins in bacteria: Bacterial IQ,extroverts and introverts

Background  

Analysis of complete microbial genomes showed that intracellular parasites and other microorganisms that inhabit stable ecological niches encode relatively primitive signaling systems, whereas environmental microorganisms typically have sophisticated systems of environmental sensing and signal transduction.     

Lipopolysaccharide-induced Notch signaling activation through JNK-dependent pathway regulates inflammatory response

Background  

Notch and TLR pathways were found to act cooperatively to activate Notch target genes and to increase the production of TLR-induced cytokines in macrophages. However, the mechanism of LPS-induced Notch activation and its role in sepsis still remains unclear.