The Human Cytomegalovirus UL116 Glycoprotein Is a Chaperone to Control gH-Based Complexes Levels on Virions

Human cytomegalovirus (HCMV) relies in large part upon the viral membrane fusion glycoprotein B and two alternative gH/gL complexes, gH/gL/gO (Trimer) and gH/gL/UL128/UL130/UL131A (Pentamer) to enter into cells. The relative amounts of Trimer and Pentamer vary among HCMV strains and contribute to differences in cell tropism. Although the viral ER resident protein UL148 has been shown to interact with gH to facilitate gO incorporation, the mechanisms that favor the assembly and maturation of one complex over another remain poorly understood. HCMV virions also contain an alternative non-disulfide bound heterodimer comprised of gH and UL116 whose function remains unknown. Here, we show that disruption of HCMV gene UL116 causes infectivity defects of ∼10-fold relative to wild-type virus and leads to reduced expression of both gH/gL complexes in virions. Furthermore, gH that is not covalently bound to other viral glycoproteins, which are readily detected in wild-type HCMV virions, become undetectable in the absence of UL116 suggesting that the gH/UL116 complex is abundant in virions. We find evidence that UL116 and UL148 interact during infection indicating that the two proteins might cooperate to regulate the abundance of HCMV gH complexes. Altogether, these results are consistent with a role of UL116 as a chaperone for gH during the assembly and maturation of gH complexes in infected cells

Moraxella catarrhalis evades neutrophil oxidative stress responses providing a safer niche for nontypeable Haemophilus influenzae

Moraxella catarrhalis and nontypeable Haemophilus influenzae (NTHi) are pathogenic bacteria frequently associated with exacerbation of chronic obstructive pulmonary disease (COPD), whose hallmark is inflammatory oxidative stress. Neutrophils produce reactive oxygen species (ROS) which can boost antimicrobial response by promoting neutrophil extracellular traps (NET) and autophagy. Here, we showed that M. catarrhalis induces less ROS and NET production in differentiated HL-60 cells compared to NTHi. It is also able to actively interfere with these responses in chemically activated cells in a phagocytosis and opsonin-independent and contact-dependent manner, possibly by engaging host immunosuppressive receptors. M. catarrhalis subverts the autophagic pathway of the phagocytic cells and survives intracellularly. It also promotes the survival of NTHi which is otherwise susceptible to the host antimicrobial arsenal. In-depth understanding of the immune evasion strategies exploited by these two human pathogens could suggest medical interventions to tackle COPD and potentially other diseases in which they co-exist

Transcriptome Analysis of Neisseria meningitidis in Human Whole Blood and Mutagenesis Studies Identify Virulence Factors Involved in Blood Survival

During infection Neisseria meningitidis (Nm) encounters multiple environments within the host, which makes rapid adaptation a crucial factor for meningococcal survival. Despite the importance of invasion into the bloodstream in the meningococcal disease process, little is known about how Nm adapts to permit survival and growth in blood. To address this, we performed a time-course transcriptome analysis using an ex vivo model of human whole blood infection. We observed that Nm alters the expression of ≈30% of ORFs of the genome and major dynamic changes were observed in the expression of transcriptional regulators, transport and binding proteins, energy metabolism, and surface-exposed virulence factors. In particular, we found that the gene encoding the regulator Fur, as well as all genes encoding iron uptake systems, were significantly up-regulated. Analysis of regulated genes encoding for surface-exposed proteins involved in Nm pathogenesis allowed us to better understand mechanisms used to circumvent host defenses. During blood infection, Nm activates genes encoding for the factor H binding proteins, fHbp and NspA, genes encoding for detoxifying enzymes such as SodC, Kat and AniA, as well as several less characterized surface-exposed proteins that might have a role in blood survival. Through mutagenesis studies of a subset of up-regulated genes we were able to identify new proteins important for survival in human blood and also to identify additional roles of previously known virulence factors in aiding survival in blood. Nm mutant strains lacking the genes encoding the hypothetical protein NMB1483 and the surface-exposed proteins NalP, Mip and NspA, the Fur regulator, the transferrin binding protein TbpB, and the L-lactate permease LctP were sensitive to killing by human blood. This increased knowledge of how Nm responds to adaptation in blood could also be helpful to develop diagnostic and therapeutic strategies to control the devastating disease cause by this microorganism

Identification of iron-activated and -repressed Fur-dependent genes by transcriptome analysis of Neisseria meningitidis group B

Iron is limiting in the human host, and bacterial pathogens respond to this environment by activating genes required for bacterial virulence. Transcriptional regulation in response to iron in Gram-negative bacteria is largely mediated by the ferric uptake regulator protein Fur, which in the presence of iron binds to a specific sequence in the promoter regions of genes under its control and acts as a repressor. Here we describe DNA microarray, computational and in vitro studies to define the Fur regulon in the human pathogen Neisseria meningitidis group B (strain MC58). After iron addition to an iron-depleted bacterial culture, 153 genes were up-regulated and 80 were down-regulated. Only 50% of the iron-regulated genes were found to contain Fur-binding consensus sequences in their promoter regions. Forty-two promoter regions were amplified and 32 of these were shown to bind Fur by gel-shift analysis. Among these genes, many of which had never been described before to be Fur-regulated, 10 were up-regulated on iron addition, demonstrating that Fur can also act as a transcriptional activator. Sequence alignment of the Fur-binding regions revealed that the N. meningitidis Fur-box encompasses the highly conserved (NATWAT)(3) motif. Cluster analysis was effective in predicting Fur-regulated genes even if computer prediction failed to identify Fur-box-like sequences in their promoter regions. Microarray-generated gene expression profiling appears to be a very effective approach to define new regulons and regulatory pathways in pathogenic bacteria