Small Hydrophobic Protein Research Articles

Transport mechanisms of SARS-CoV-E viroporin in calcium solutions: Lipid-dependent Anomalous Mole Fraction Effect and regulation of pore conductance

The envelope protein E of the SARS-CoV coronavirus is an archetype of viroporin. It is a small hydrophobic protein displaying ion channel activity that has proven highly relevant in virus-host interaction and virulence. Ion transport through E channel was shown to alter Ca2+ homeostasis in the cell and trigger inflammation processes. Here, we study transport properties of the E viroporin in mixed solutions of potassium and calcium chloride that contain a fixed total concentration (mole fraction experiments). The channel is reconstituted in planar membranes of different lipid compositions, including a lipid mixture that mimics the endoplasmic reticulum-Golgi intermediate compartment (ERGIC) membrane where the virus localizes within the cell. We find that the E ion conductance changes non-monotonically with the total ionic concentration displaying an Anomalous Mole Fraction Effect (AMFE) only when charged lipids are present in the membrane. We also observe that E channel insertion in ERGIC-mimic membranes – including lipid with intrinsic negative curvature – enhances ion permeation at physiological concentrations of pure CaCl2 or KCl solutions, with a preferential transport of Ca2+ in mixed KCl-CaCl2 solutions. Altogether, our findings demonstrate that the presence of calcium modulates the transport properties of the E channel by interacting preferentially with charged lipids through different mechanisms including direct Coulombic interactions and possibly inducing changes in membrane morphology.

Subcellular Journey of Rare Cold Inducible 2 Protein in Plant Under Stressful Condition.

Rare cold inducible 2 (RCI2) proteins are small hydrophobic membrane proteins in plants, and it has been widely reported that RCI2 expressions are dramatically induced by salt, cold, and drought stresses in many species. The RCI2 proteins have been shown to regulate plasma membrane (PM) potential and enhance abiotic stress tolerance when over-expressed in plants. RCI2 protein structures contain two transmembrane domains that are thought to be PM intrinsic proteins and have been observed at the PM and endomembranes. However, cellular trafficking of RCI2s are not fully understood. In this review, we discussed (i) general properties of RCI2s characterized in many species, (ii) the uses of RCI2s as a tracer in live cell imaging analyses and when they are fused to fluorescence proteins during investigations into vesicle trafficking, and (iii) RCI2 functionalities such as their involvement in rapid diffusion, endocytosis, and protein interactions. Consequently, the connection between physiological characteristics of RCI2s and traffic of RCI2s interacting membrane proteins might be helpful to understand role of RCI2s contributing abiotic stresses tolerance.

Expression and Regulation of Aquaporins in Pregnancy Complications and Reproductive Dysfunctions.

Aquaporins (AQPs), small hydrophobic integral membrane proteins, mediate rapid transport of water and small solutes. The abnormal expressions of AQPs are associated with pregnancy complications and reproductive dysfunctions, including preeclampsia, gestational diabetes mellitus, tubal ectopic pregnancy, intrahepatic cholestasis of pregnancy, preterm birth, chorioamnionitis, polyhydramnios, and oligohydramnios, thus resulting in adverse pregnancy outcomes. This review explains the alterations of AQPs in pregnancy complications and reproductive dysfunctions and summarizes the molecular mechanisms involved in the regulations of AQPs by drugs such as oxytocin, polychlorinated biphenyls, all-trans-retinoic acid, salvia miltiorrhiza, and insulin, or other factors such as oxygen and osmotic pressure. All the research provides evidence that AQPs could be the new therapeutic targets of pregnancy-related diseases.

Functional Characterization of Circulating Mumps Viruses with Stop Codon Mutations in the Small Hydrophobic Protein.

Between 2015 and 2017, routine molecular surveillance in the United States detected multiple mumps viruses (MuVs) with mutations in the small hydrophobic (SH) gene compared to a reference virus of the same genotype. These mutations include an unusual pattern of uracil-to-cytosine hypermutations and other mutations resulting in the generation of premature stop codons or disruption of the canonical stop codon. The mumps virus SH protein may serve as a virulence factor, based on evidence that it inhibits apoptosis and innate immune signaling in vitro and that recombinant viruses that do not express the SH protein are attenuated in an animal model. In this study, mumps viruses bearing variant SH sequences were isolated from contemporary outbreak samples to evaluate the impact of the observed mutations on SH protein function. All isolates with variant SH sequences replicated in interferon-competent cells with no evidence of attenuation. Furthermore, all SH-variant viruses retained the ability to abrogate induction of NF-κB-mediated innate immune signaling in infected cells. Ectopic expression of variant mumps SH genes is consistent with findings from infection experiments, indicating that the observed abrogation of signaling was not mediated by other viral factors that may modulate innate immune signaling. Molecular surveillance is an important public health tool for monitoring the diversity of circulating mumps viruses and can provide insights into determinants of disease. These findings, in turn, will inform studies employing reverse genetics to elucidate the specific mechanisms of MuV pathogenesis and potential impacts of observed sequence variants on infectivity, fitness, and virulence.IMPORTANCE Mumps virus (MuV) outbreaks occur in the United States despite high coverage with measles, mumps, rubella (MMR) vaccine. Routine genotyping of laboratory-confirmed mumps cases has been practiced in the United States since 2006 to enhance mumps surveillance. This study reports the detection of unusual mutations in the small hydrophobic (SH) protein of contemporary laboratory-confirmed mumps cases and is the first to describe the impact of such mutations on SH protein function. These mutations are predicted to profoundly alter the amino acid sequence of the SH protein, which has been shown to antagonize host innate immune responses; however, they were neither associated with defects in virus replication nor attenuated protein function in vitro, consistent with detection in clinical specimens. A better understanding of the forces governing mumps virus sequence diversity and of the functional consequences of mutations in viral proteins is important for maintaining robust capacity for mumps detection and disease control.

GBP5 Is an Interferon-Induced Inhibitor of Respiratory Syncytial Virus.

Guanylate binding protein 5 (GBP5) belongs to the GTPase subfamily, which is mainly induced by interferon gamma (IFN-γ) and is involved in many important cellular processes, including inflammasome activation and innate immunity against a wide variety of microbial pathogens. However, it is unknown whether GBP5 inhibits respiratory syncytial virus (RSV) infection. In this study, we identified GBP5 as an effector of the anti-RSV activity of IFN-γ and found that in children, the weaker immune response, especially the weaker IFN-γ response and the decreased GBP5 expression, leads to RSV susceptibility. Furthermore, we revealed that GBP5 reduced the cell-associated levels of the RSV small hydrophobic (SH) protein, which was identified as a viroporin. In contrast, overexpression of the SH protein rescued RSV replication in the presence of GBP5. The GBP5-induced decrease in intracellular SH protein levels is because GBP5 promotes the release of the SH protein into the cell culture. Moreover, the GBP5 C583A mutants with changes at the C terminus or the GBP5 ΔC mutant lacking the C-terminal region, which impairs GBP5 localization in the Golgi, could not inhibit RSV infection, whereas the GTPase-defective GBP5 maintained RSV inhibition, suggesting that Golgi localization but not the GTPase activity of GBP5 is required for RSV inhibition. Interestingly, we found that RSV infection or RSV G protein downregulates GBP5 expression by upregulating DZIP3, an E3 ligase, which induces GBP5 degradation through the K48 ubiquitination and proteasomal pathways. Thus, this study reveals a complicated interplay between host restrictive factor GBP5 and RSV infection and provides important information for understanding the pathogenesis of RSV.IMPORTANCE RSV is a highly contagious virus that causes multiple infections in infants within their first year of life. It can also easily cause infection in elderly or immunocompromised individuals, suggesting that individual differences in immunity play an important role in RSV infection. Therefore, exploring the pathogenic mechanisms of RSV and identifying essential genes which inhibit RSV infection are necessary to develop an effective strategy to control RSV infection. Here, we report that the IFN-inducible gene GBP5 potently inhibits RSV replication by reducing the cell-associated levels of the RSV small hydrophobic (SH) protein, which is a viroporin. In contrast, the RSV G protein was shown to upregulate the expression of the DZIP3 protein, an E3 ligase that degrades GBP5 through the proteasomal pathway. Our study provides important information for the understanding of the pathogenic mechanisms of RSV and host immunity as well as the complicated interplay between the virus and host.

Uncovering small membrane proteins in pathogenic bacteria: Regulatory functions and therapeutic potential.

Bacterial small proteins (below 50 amino acids) encoded by small open reading frames (sORFs) are recognized as an emerging class of functional molecules that have been largely overlooked in the past. While some were uncovered serendipitously, global approaches have recently been developed to detect these sORFs. A large portion of small proteins appears to be hydrophobic and located in the bacterial membrane. In the present review, we describe functional small hydrophobic proteins discovered in pathogenic bacteria and report recent advances in the discovery of additional ones. Small membrane proteins contribute to bacterial adaptation to changing environments and often appear to be implicated in negative feedback regulation loops by modulating the function or stability of larger membrane proteins. A subset of these proteins belongs to toxin-antitoxin modules. We highlight the features of characterized hydrophobic small proteins that may pave the way for identification of the functional small proteins among novel sORFs discovered. Besides providing new insights into bacterial pathogenesis, identification of naturally occurring small hydrophobic proteins of pathogenic bacteria can lead to new therapeutic interventions, as recently shown with the development of synthetic peptides derived from natural small proteins that display antibacterial or antivirulence properties.

Evaluation of the protective potential of antibody and T cell responses elicited by a novel preventative vaccine towards respiratory syncytial virus small hydrophobic protein

ABSTRACT The small hydrophobic (SH) glycoprotein of human respiratory syncytial virus (RSV) is a transmembrane protein that is poorly accessible by antibodies on the virion but has an ectodomain (SHe) that is accessible and expressed on infected cells. The SHe from RSV strain A has been formulated in DPX, a unique delivery platform containing an adjuvant, and is being evaluated as an RSV vaccine candidate. The proposed mechanism of protection is the immune-mediated clearance of infected cells rather than neutralization of the virion. Our phase I clinical trial data clearly showed that vaccination resulted in robust antibody responses, but it was unclear if these immune responses have any correlation to immune responses to natural infection with RSV. Therefore, we embarked on this study to examine these immune responses in older adults with confirmed RSV infection. We compared vaccine-induced (DPX-RSV(A)) immune responses from participants in a Phase 1 clinical trial to paired acute and convalescent titers from older adults with symptomatic laboratory-confirmed RSV infection. Serum samples were tested for anti-SHe IgG titers and the isotypes determined. T cell responses were evaluated by IFN-γ ELISPOT. Anti-SHe titers were detected in 8 of 42 (19%) in the acute phase and 16 of 42 (38%) of convalescent serum samples. IgG1, IgG3, and IgA were the prevalent isotypes generated by both vaccination and infection. Antigen-specific T cell responses were detected in 9 of 16 (56%) of vaccinated participants. Depletion of CD4+ but not CD8+ T cells abrogated the IFN-γ ELISPOT response supporting the involvement of CD4+ T cells in the immune response to vaccination. The data showed that an immune response like that induced by DPX-RSV(A) could be seen in a subset of participants with confirmed RSV infection. These findings show that older adults with clinically significant infection as well as vaccinated adults generate a humoral response to SHe. The induction of both SHe-specific antibody and cellular responses support further clinical development of the DPX-RSV(A) vaccine.

Genetic characterization of mumps viruses associated with the resurgence of mumps in the United States: 2015–2017

Despite high coverage with measles, mumps, and rubella vaccine in the United States, outbreaks of mumps occur in close contact settings such as schools, colleges, and camps. Starting in late 2015, outbreaks were reported from several universities, and by the end of 2017, greater than 13,800 cases had been reported nation-wide. In 2013, the CDC and the Association of Public Health Laboratories contracted four Vaccine Preventable Diseases Reference Centers (VPD-RCs) to perform real-time reverse transcription PCR (RT-qPCR) to detect mumps RNA in clinical samples and to determine the genotype. Twelve genotypes of mumps virus are currently recognized by the World Health Organization, and the standard protocol for genotyping requires sequencing the entire gene coding for the small hydrophobic (SH) protein. Phylogenetic analysis of the 1862 mumps samples genotyped from 2015 through 2017 showed that the overall diversity of genotypes detected was low. Only 0.8 % of the sequences were identified as genotypes C, H, J, or K, and 0.5 % were identified as vaccine strains in genotypes A or N, while most sequences (98.7 %) were genotype G. The majority of the genotype G sequences could be included into one of two large groups with identical SH sequences. Within genotype G, a small number of phylogenetically significant outlier sequences were associated with epidemiologically distinct chains of transmission. These results demonstrate that molecular and epidemiologic data can be used to track transmission pathways of mumps virus; however, the limited diversity of the SH sequences may be insufficient for resolving transmission in all outbreaks.

Parainfluenza Virus 5 Infection in Neurological Disease and Encephalitis of Cattle.

The etiology of viral encephalitis in cattle often remains unresolved, posing a potential risk for animal and human health. In metagenomics studies of cattle with bovine non-suppurative encephalitis, parainfluenza virus 5 (PIV5) was identified in three brain samples. Interestingly, in two of these animals, bovine herpesvirus 6 and bovine astrovirus CH13 were additionally found. We investigated the role of PIV5 in bovine non-suppurative encephalitis and further characterized the three cases. With traditional sequencing methods, we completed the three PIV5 genomes, which were compared to one another. However, in comparison to already described PIV5 strains, unique features were revealed, like an 81 nucleotide longer open reading frame encoding the small hydrophobic (SH) protein. With in situ techniques, we demonstrated PIV5 antigen and RNA in one animal and found a broad cell tropism of PIV5 in the brain. Comparative quantitative analyses revealed a high viral load of PIV5 in the in situ positive animal and therefore, we propose that PIV5 was probably the cause of the disease. With this study, we clearly show that PIV5 is capable of naturally infecting different brain cell types in cattle in vivo and therefore it is a probable cause of encephalitis and neurological disease in cattle.

Small hydrophobic viral proteins involved in intercellular movement of diverse plant virus genomes.

Most plant viruses code for movement proteins (MPs) targeting plasmodesmata to enable cell-to-cell and systemic spread in infected plants. Small membrane-embedded MPs have been first identified in two viral transport gene modules, triple gene block (TGB) coding for an RNA-binding helicase TGB1 and two small hydrophobic proteins TGB2 and TGB3 and double gene block (DGB) encoding two small polypeptides representing an RNA-binding protein and a membrane protein. These findings indicated that movement gene modules composed of two or more cistrons may encode the nucleic acid-binding protein and at least one membrane-bound movement protein. The same rule was revealed for small DNA-containing plant viruses, namely, viruses belonging to genus Mastrevirus (family Geminiviridae) and the family Nanoviridae. In multi-component transport modules the nucleic acid-binding MP can be viral capsid protein(s), as in RNA-containing viruses of the families Closteroviridae and Potyviridae. However, membrane proteins are always found among MPs of these multicomponent viral transport systems. Moreover, it was found that small membrane MPs encoded by many viruses can be involved in coupling viral replication and cell-to-cell movement. Currently, the studies of evolutionary origin and functioning of small membrane MPs is regarded as an important pre-requisite for understanding of the evolution of the existing plant virus transport systems. This paper represents the first comprehensive review which describes the whole diversity of small membrane MPs and presents the current views on their role in plant virus movement.

Determining Immune and miRNA Biomarkers Related to Respiratory Syncytial Virus (RSV) Vaccine Types.

Respiratory Syncytial Virus (RSV) causes serious respiratory tract illness and substantial morbidity and some mortality in populations at the extremes of age, i.e., infants, young children, and the elderly. To date, RSV vaccine development has been unsuccessful, a feature linked to the lack of biomarkers available to assess the safety and efficacy of RSV vaccine candidates. We examined microRNAs (miR) as potential biomarkers for different types of RSV vaccine candidates. In this study, mice were vaccinated with a live attenuated RSV candidate that lacks the small hydrophobic (SH) and attachment (G) proteins (CP52), an RSV G protein microparticle (GA2-MP) vaccine, a formalin-inactivated RSV (FI-RSV) vaccine or were mock-treated. Several immunological endpoints and miR expression profiles were determined in mouse serum and bronchoalveolar lavage (BAL) following vaccine priming, boost, and RSV challenge. We identified miRs that were linked with immunological parameters of disease and protection. We show that miRs are potential biomarkers providing valuable insights for vaccine development.

Affinity-Triggered Assemblies Based on a Designed Peptide-Peptide Affinity Pair.

Affinity-triggered assemblies rely on affinity interactions as the driving force to assemble physically crosslinked networks. WW domains are small hydrophobic proteins binding to proline-rich peptides that are typically produced in the insoluble form. Previous works attempted the biological production of the full WW domain in tandem to generate multivalent components for affinity-triggered hydrogels. In this work, an alternative approach is followed by engineering a 13-mer minimal version of the WW domain that retains the ability to bind to target proline-rich peptides. Both ligand and target peptides are produced chemically and conjugated to multivalent polyethylene glycol, yielding two components. Upon mixing together, they form soft biocompatible affinity-triggered assemblies, stable in stem cell culture media, and display mechanical properties in the same order of magnitude as for those hydrogels formed with the full WW protein in tandem.

Potential Viroporin Candidates From Pathogenic Viruses Using Bacteria-Based Bioassays

Viroporins are a family of small hydrophobic proteins found in many enveloped viruses that are capable of ion transport. Building upon the ability to inhibit influenza by blocking its archetypical M2 H+ channel, as a family, viroporins may represent a viable target to curb viral infectivity. To this end, using three bacterial assays we analyzed six small hydrophobic proteins from biomedically important viruses as potential viroporin candidates. Our results indicate that Eastern equine encephalitis virus 6k, West Nile virus MgM, Dengue virus 2k, Dengue virus P1, Variola virus gp170, and Variola virus gp151 proteins all exhibit channel activity in the bacterial assays, and as such may be considered viroporin candidates. It is clear that more studies, such as patch clamping, will be needed to characterize the ionic conductivities of these proteins. However, our approach presents a rapid procedure to analyze open reading frames in other viruses, yielding new viroporin candidates for future detailed investigation. Finally, if conductivity is proven vital to their cognate viruses, the bio-assays presented herein afford a simple approach to screen for new channel blockers.

Type I Toxin-Antitoxin Systems in Clostridia.

Type I toxin-antitoxin (TA) modules are abundant in both bacterial plasmids and chromosomes and usually encode a small hydrophobic toxic protein and an antisense RNA acting as an antitoxin. The RNA antitoxin neutralizes toxin mRNA by inhibiting its translation and/or promoting its degradation. This review summarizes our current knowledge of the type I TA modules identified in Clostridia species focusing on the recent findings in the human pathogen Clostridium difficile. More than ten functional type I TA modules have been identified in the genome of this emerging enteropathogen that could potentially contribute to its fitness and success inside the host. Despite the absence of sequence homology, the comparison of these newly identified type I TA modules with previously studied systems in other Gram-positive bacteria, i.e., Bacillus subtilis and Staphylococcus aureus, revealed some important common traits. These include the conservation of characteristic sequence features for small hydrophobic toxic proteins, the localization of several type I TA within prophage or prophage-like regions and strong connections with stress response. Potential functions in the stabilization of genome regions, adaptations to stress conditions and interactions with CRISPR-Cas defence system, as well as promising applications of TA for genome-editing and antimicrobial developments are discussed.

Characterization of an OrtT-like toxin of Salmonella enterica serovar Houten.

The Escherichia coli GhoT/GhoS system is a type V toxin-antitoxin system in which the antitoxin GhoS cleaves the GhoT mRNA, controlling its translation. GhoT is a small hydrophobic protein that damages bacterial membranes. OrtT is a GhoT-like toxin, but it apparently lacks a corresponding antitoxin and serves a different physiologic role. Using a profile hidden Markov model approach, a Salmonella enterica serovar Houten genome was screened to obtain homologs of GhoT/OrtT. We only found one protein (referred to here as OrtT-Sal) that shared more sequence identity with OrtT than GhoT. The chromosomal region around the coding sequence of OrtT-Sal suggests that it is an orphan toxin and can be under RpoH activation. To study OrtT-Sal, we chemically synthesized and expressed in E. coli the whole toxin and its N- and C-terminal regions (OrtT-Sal1-29 and OrtT-Sal29-57, respectively). Our findings have shown that the overproduction of the polypeptides resulted in severe growth inhibition and cell lysis. Using circular dichroism, we found that OrtT-Sal, OrtT-Sal1-29, and OrtT-Sal29-57 form an alpha-helical structure in the presence of SDS micelles or TFE. Finally, using carboxyfluorescein-loaded lipid vesicles, we determined that the polypeptides damage lipid membrane directly.

Human metapneumovirus activates NOD-like receptor protein 3 inflammasome via its small hydrophobic protein which plays a detrimental role during infection in mice.

NOD-like receptor protein 3 (NLRP3) inflammasome activation triggers caspase-1 activation-induced maturation of interleukin (IL)-1β and IL-18 and therefore is important for the development of the host defense against various RNA viral diseases. However, the implication of this protein complex in human metapneumovirus (HMPV) disease has not been fully studied. Herein, we report that NLRP3 inflammasome plays a detrimental role during HMPV infection because NLRP3 inflammasome inhibition protected mice from mortality and reduced weight loss and inflammation without impacting viral replication. We also demonstrate that NLRP3 inflammasome exerts its deleterious effect via IL-1β production since we observed reduced mortality, weight loss and inflammation in IL-1β-deficient (IL-1β-/-) mice, as compared to wild-type animals during HMPV infection. Moreover, the effect on these evaluated parameters was not different in IL-1β-/- and wild-type mice treated with an NLRP3 inflammasome inhibitor. The production of IL-1β was also abrogated in bone marrow derived macrophages deficient for NLRP3. Finally, we show that small hydrophobic protein-deleted recombinant HMPV (HMPV ΔSH) failed to activate caspase-1, which is responsible for IL-1β cleavage and maturation. Furthermore, HMPV ΔSH-infected mice had less weight loss, showed no mortality and reduced inflammation, as compared to wild-type HMPV-infected mice. Thus, NLRP3 inflammasome activation seems to be triggered by HMPV SH protein in HMPV disease. In summary, once activated by the HMPV SH protein, NLRP3 inflammasome promotes the maturation of IL-1β, which exacerbates HMPV-induced inflammation. Therefore, the blockade of IL-1β production by using NLRP3 inflammasome inhibitors might be a novel potential strategy for the therapy and prevention of HMPV infection.

NaCl-induced CsRCI2E and CsRCI2F interact with aquaporin CsPIP2;1 to reduce water transport in Camelina sativa L.

Rare cold-inducible 2 (RCI2) proteins are small hydrophobic proteins that are known to be localized in cellular membranes. The function of RCI2 proteins has been reported to be associated with low-temperature, salt, and drought stress tolerances as a membrane potential regulator; however, the specific functions are still unknown. The PIP2 (plasma membrane intrinsic protein 2) aquaporins are proteins that transport water and small solutes into the cell. The expression and activity of PIP2 proteins, like RCI2, are also related to salt- and drought-stress tolerance. In this study, we identified novel protein interactions between RCI2 and PIP2; 1, including protein accumulation changes in the bioenergy crop Camelina sativa L. under various NaCl stress conditions. Accumulation of both CsRCI2E and CsRCI2F proteins increased with NaCl stress; however, to differing levels depending on the NaCl stress intensity. A co-immunoprecipitation test revealed interaction between CsRCI2E-CsPIP2 and CsRCI2F-CsPIP2. Moreover, co-expression of the four CsRCI2 proteins with CsPIP2; 1 in Xenopus laevis oocytes reduced water transport activity. Furthermore, the abundance of CsPIP2; 1 protein was decreased under CsRCI2E and CsRCI2F co-expression. These results suggest that NaCl-induced expression of CsRCI2E and CsRCI2F contributes to the regulation of CsPIP2; 1.

Genetic characteristics of mumps virus isolated from one outbreak in Henan province in 2016

Objective To analyze the genetic characteristics of mumps virus (MuV) isolated from an outbreak in Henan province in 2016 and to provide reference data for the control of mumps in Henan province. Methods Swab specimens were collected from a mumps outbreak in Henan province in 2016. One virus strain was selected for whole genome sequencing after virus isolation and genotyping. MEGA7.0 software was used to construct phylogenetic tree, to calculate P-distance and to analyze the characteristics of genes encoding small hydrophobic protein (SH), fusion protein (F) and hemagglutinin neuraminidase (HN). Results Five swab specimens were collected in the outbreak and five MuV strains were successfully obtained. Phylogenetic analysis showed that all of them belonged to genotype F with a P-distance of 0.047 (0.046-0.049) with the reference strain. The P-distance among the five MuV strains was 0.001 (0-0.003). Results of the whole genome sequencing of one MuV strain showed that the P-distance with the reference virus was 0.053 (0.018-0.072) and the smallest P-distance with genotype F was 0.018. Compared with the vaccine strain (HQ416907.1), the amino acid at 28-30 site of SH protein was IML. The 91, 195 and 383 amino acid sites in F protein were related to the pathogenicity of MuV, but no mutations were found in these sites. Some mutations were found in HN protein, such as P354Q, E356D and K464N, which were associated with the immunological characteristics of MuV. Conclusions MuV strains isolated from the outbreak in Henan province in 2016 belonged to genotype F. The genetic characteristics of these MuV strains were similar to those prevalent in other provinces of China. Key words: Mumps virus; Genetic characteristics; Analysis

Mechanisms of Resistance to the Contact-Dependent Bacteriocin CdzC/D in Caulobacter crescentus.

The Cdz bacteriocin system allows the aquatic oligotrophic bacterium Caulobacter crescentus to kill closely related species in a contact-dependent manner. The toxin, which aggregates on the surfaces of producer cells, is composed of two small hydrophobic proteins, CdzC and CdzD, each bearing an extended glycine-zipper motif, that together induce inner membrane depolarization and kill target cells. To further characterize the mechanism of Cdz delivery and toxicity, we screened for mutations that render a target strain resistant to Cdz-mediated killing. These mutations mapped to four loci, including a TonB-dependent receptor, a three-gene operon (named zerRAB for zipper envelope resistance), and perA (for pentapeptide envelope resistance). Mutations in the zerRAB locus led to its overproduction and to potential changes in cell envelope composition, which may diminish the susceptibility of cells to Cdz toxins. The perA gene is also required to maintain a normal cell envelope, but our screen identified mutations that confer resistance to Cdz toxins without substantially affecting the cell envelope functions of PerA. We demonstrate that PerA, which encodes a pentapeptide repeat protein predicted to form a quadrilateral β-helix, localizes primarily to the outer membrane of cells, where it may serve as a receptor for the Cdz toxins. Collectively, these results provide new insights into the function and mechanisms of an atypical, contact-dependent bacteriocin system.IMPORTANCE Bacteriocins are commonly used by bacteria to kill neighboring cells that compete for resources. Although most bacteriocins are secreted, the aquatic, oligotrophic bacterium Caulobacter crescentus produces a two-peptide bacteriocin, CdzC/D, that remains attached to the outer membranes of cells, enabling contact-dependent killing of cells lacking the immunity protein CdzI. The receptor for CdzC/D has not previously been reported. Here, we describe a genetic screen for mutations that confer resistance to CdzC/D. One locus identified, perA, encodes a pentapeptide repeat protein that resides in the outer membrane of target cells, where it may act as the direct receptor for CdzC/D. Collectively, our results provide new insight into bacteriocin function and diversity.

The small hydrophobic (SH) gene of North American turkey AMPV-C does not attenuate nor modify host tropism in recombinant European duck AMPV-C

Subgroup C Avian Metapneumoviruses (AMPV-C) has two lineages, one mostly in turkeys and one mostly in ducks. To investigate the molecular basis of AMPV-C host tropism, a reverse genetics system for a duck AMPV-C virus was developed. A recombinant copy and a recombinant virus in which the SH protein had been exchanged for that of a turkey AMPV-C were rescued. No change in cytopathogenic effect or replication profile in vitro were observed for either virus compared to the wild type. In SPF Muscovy ducks the wild type and its recombinant copy were equally pathogenic. Exchanging the SH in the recombinant copy produced the same results.In SPF turkeys, neither recombinant virus was pathogenic, although both showed a low level of replication. Thus, from the current model, it appears that AMPV-C SH proteins derived from the different species are compatible and that turkey SH does not affect duck AMPV-C pathogenicity.