TLR4 Asp299Gly and Thr399Ile Polymorphisms: No Impact on Human Immune Responsiveness to LPS or Respiratory Syncytial Virus

A broad variety of natural environmental stimuli, genotypic influences and timing all contribute to expression of protective versus maladaptive immune responses and the resulting clinical outcomes in humans. The role of commonly co-segregating Toll-like receptor 4 (TLR4) non-synonymous single nucleotide polymorphisms Asp299Gly and Thr399Ile in this process remains highly controversial. Moreover, what differential impact these polymorphisms might have in at risk populations with respiratory dysfunction, such as current asthma or a history of infantile bronchiolitis, has never been examined. Here we determine the importance of these polymorphisms in modulating LPS and respiratory syncytial virus (RSV)--driven cytokine responses. We focus on both healthy children and those with clinically relevant respiratory dysfunction.To elucidate the impact of TLR4 Asp299Gly and Thr399Ile on cytokine production, we assessed multiple immune parameters in over 200 pediatric subjects aged 7-9. Genotyping was followed by quantification of pro- and anti-inflammatory cytokine responses by fresh peripheral blood mononuclear cells upon acute exposure to LPS or RSV.In contrast to early reports, neither SNP influenced immune responses evoked by LPS exposure or RSV infection, as measured by the intermediate phenotype of pro- and anti-inflammatory cytokine responses to these ubiquitous agents. There is no evidence of altered sensitivity in populations with "at risk" clinical phenotypes.Genomic medicine seeks to inform clinical practice. Determination of the TLR4 Asp299Gly/Thr399Ile haplotype is of no clinical benefit in predicting the nature or intensity of cytokine production in children whether currently healthy or among specific at-risk groups characterized by prior infantile broncholitis or current asthma

Endogenous laminin is required for human airway smooth muscle cell maturation

BACKGROUND: Airway smooth muscle (ASM) contraction underlies acute bronchospasm in asthma. ASM cells can switch between a synthetic-proliferative phenotype and a contractile phenotype. While the effects of extracellular matrix (ECM) components on modulation of ASM cells to a synthetic phenotype have been reported, the role of ECM components on maturation of ASM cells to a contractile phenotype in adult lung is unclear. As both changes in ECM components and accumulation of contractile ASM are features of airway wall remodelling in asthma, we examined the role of the ECM protein, laminin, in the maturation of contractile phenotype in human ASM cells. METHODS: Human ASM cells were made senescence-resistant by stable expression of human telomerase reverse transcriptase. Maturation to a contractile phenotype was induced by 7-day serum deprivation, as assessed by immunoblotting for desmin and calponin. The role of laminin on ASM maturation was investigated by comparing the effects of exogenous laminin coated on culture plates, and of soluble laminin peptide competitors. Endogenous expression of laminin chains during ASM maturation was also measured. RESULTS: Myocyte binding to endogenously expressed laminin was required for ASM phenotype maturation, as laminin competing peptides (YIGSR or GRGDSP) significantly reduced desmin and calponin protein accumulation that otherwise occurs with prolonged serum deprivation. Coating of plastic cell culture dishes with different purified laminin preparations was not sufficient to further promote accumulation of desmin or calponin during 7-day serum deprivation. Expression of α2, β1 and γ1 laminin chains by ASM cells was specifically up-regulated during myocyte maturation, suggesting a key role for laminin-2 in the development of the contractile phenotype. CONCLUSION: While earlier reports suggest exogenously applied laminin slows the spontaneous modulation of ASM to a synthetic phenotype, we show for the first time that endogenously expressed laminin is required for ASM maturation to the contractile phenotype. As endogenously expressed laminin chains α2, β1 and γ1 are uniquely increased during myocyte maturation, these laminin chains may be key in this process. Thus, human ASM maturation appears to involve regulated endogenous expression of a select set of laminin chains that are essential for accumulation of contractile phenotype myocytes

Detection of solar dimming and brightening effects on Northern Hemisphere river flow

Anthropogenic aerosols in the atmosphere have the potential to affect regional-scale land hydrology through solar dimming1, 2. Increased aerosol loading may have reduced historical surface evaporation over some locations3, but the magnitude and extent of this effect is uncertain. Any reduction in evaporation due to historical solar dimming may have resulted in an increase in river flow. Here we formally detect and quantify the historical effect of changing aerosol concentrations, via solar radiation, on observed river flow over the heavily industrialized, northern extra-tropics. We use a state-of-the-art estimate of twentieth century surface meteorology4 as input data for a detailed land surface model5, and show that the simulations capture the observed strong inter-annual variability in runoff in response to climatic fluctuations. Using statistical techniques, we identify a detectable aerosol signal in the observed river flow both over the combined region, and over individual river basins in Europe and North America. We estimate that solar dimming due to rising aerosol concentrations in the atmosphere around 1980 led to an increase in river runoff by up to 25% in the most heavily polluted regions in Europe. We propose that, conversely, these regions may experience reduced freshwater availability in the future, as air quality improvements are set to lower aerosol loading and solar dimming

Investigating the SecY plug movement at the SecYEG translocation channel

Protein translocation occurs across the energy-conserving bacterial membrane at the SecYEG channel. The crystal structure of the channel has revealed a possible mechanism for gating and opening. This study evaluates the plug hypothesis using cysteine crosslink experiments in combination with various allelic forms of the Sec complex. The results demonstrate that the SecY plug domain moves away from the center of the channel toward SecE during polypeptide translocation, and further show that the translocation-enhancing prlA3 mutation and SecG subunit change the properties of channel gating. Locking the plug in the open state preactivates the Sec complex, and a super-active translocase can be created when combined with the prlA4 mutation located in the pore of the channel. Dimerization of the Sec complex, which is essential for translocase activity, relocates the plug toward the open position. We propose that oligomerization may result in SecYEG cooperative interactions important to prime the translocon function

The novel coactivator C1 (HCF) coordinates multiprotein enhancer formation and mediates transcription activation by GABP

Transcription of the herpes simplex virus 1 (HSV–1) immediate early (IE) genes is determined by multiprotein enhancer complexes. The core enhancer assembly requires the interactions of the POU-homeodomain protein Oct–1, the viral transactivator αTIF and the cellular factor C1 (HCF). In this context, the C1 factor interacts with each protein to assemble the stable enhancer complex. In addition, the IE enhancer cores contain adjacent binding sites for other cellular transcription factors such as Sp1 and GA-binding protein (GABP). In this study, a direct interaction of the C1 factor with GABP is demonstrated, defining the C1 factor as the critical coordinator of the enhancer complex assembly. In addition, mutations that reduce the GABP transactivation potential also impair the C1–GABP interaction, indicating that the C1 factor functions as a novel coactivator of GABP-mediated transcription. The interaction and coordinated assembly of the enhancer proteins by the C1 factor may be critical for the regulation of the HSV lytic–latent cycle