Virus Encoded MHC-Like Decoys Diversify the Inhibitory KIR Repertoire

Natural killer (NK) cells are circulating lymphocytes that play an important role in the control of viral infections and tumors. Their functions are regulated by several activating and inhibitory receptors. A subset of these receptors in human NK cells are the killer immunoglobulin-like receptors (KIRs), which interact with the highly polymorphic MHC class I molecules. One important function of NK cells is to detect cells that have down-regulated MHC expression (missing-self). Because MHC molecules have non polymorphic regions, their expression could have been monitored with a limited set of monomorphic receptors. Surprisingly, the KIR family has a remarkable genetic diversity, the function of which remains poorly understood. The mouse cytomegalovirus (MCMV) is able to evade NK cell responses by coding “decoy” molecules that mimic MHC class I. This interaction was suggested to have driven the evolution of novel NK cell receptors. Inspired by the MCMV system, we develop an agent-based model of a host population infected with viruses that are able to evolve MHC down-regulation and decoy molecules. Our simulations show that specific recognition of MHC class I molecules by inhibitory KIRs provides excellent protection against viruses evolving decoys, and that the diversity of inhibitory KIRs will subsequently evolve as a result of the required discrimination between host MHC molecules and decoy molecules.     

Autocrine ligand binding to cell receptors. Mathematical analysis of competition by solution "decoys".

Autocrine ligands have been demonstrated to regulate cell proliferation, cell adhesion, and cell migration in a number of different systems and are believed to be one of the underlying causes of malignant cell transformation. Binding of these ligands to their cellular receptors can be compromised by diffusive transport of ligand away from the secreting cell. Exogenous addition of antibodies or solution receptors capable of competing with cellular receptors for these autocrine ligands has been proposed as a means of inhibiting autocrine-stimulated cell behavioral responses. Such "decoys" complicate cellular binding by offering alternative binding targets, which may also be capable of aiding or abating transport of the ligand away from the cell surface. We present a mathematical model incorporating autocrine ligand production and the presence of competing cellular and solution receptors. We elucidate effects of key system parameters including ligand diffusion rate, binding rate constants, cell density, and secretion rate on the ability of solution receptors to inhibit cellular receptor binding. Both plated and suspension cell systems are considered. An approximate analytical expression relating the key parameters to the critical concentration of solution "decoys" required for inhibition is derived and compared to the numerical calculations. We find that in order to achieve essentially complete inhibition of surface receptor binding, the concentration of decoys may need to be as much as four to eight orders of magnitude greater than the equilibrium disociation constant for ligand binding to surface receptors.     

Tuning inflammation and immunity by chemokine sequestration: decoys and more

A set of chemokine receptors are structurally unable to elicit migration or conventional signalling responses after ligand engagement. These 'silent' (non-signalling) chemokine receptors regulate inflammatory and immune reactions in different ways, including by acting as decoys and scavengers. Chemokine decoy receptors recognize distinct and complementary sets of ligands and are strategically expressed in different cellular contexts. Importantly, viruses and parasites have evolved multiple strategies to elude chemokines, including the expression of decoy receptors. So, decoy receptors for chemokines represent a general strategy to tune, shape and temper innate and adaptive immunity.     

Targeting host lipid flows: Exploring new antiviral and antibiotic strategies

Bacteria and viruses pose serious challenges for humans because they evolve continuously. Despite ongoing efforts, antiviral drugs to treat many of the most troubling viruses have not been approved yet. The recent launch of new antimicrobials is generating hope as more and more pathogens around the world become resistant to available drugs. But extra effort is still needed. One of the current strategies for antiviral and antibiotic drug development is the search for host cellular pathways used by many different pathogens. For example, many viruses and bacteria alter lipid synthesis and transport to build their own organelles inside infected cells. The characterization of these interactions will be fundamental to identify new targets for antiviral and antibiotic drug development. This review discusses how viruses and bacteria subvert cell machineries for lipid synthesis and transport and summarises the most promising compounds that interfere with these pathways.     

Cytokine involvement in viral permissiveness and the progression of HIV disease

Many viruses have evolved novel means of exploiting host defense mechanisms for their own survival. This exploitation may be best exemplified by the interrelationships between certain viruses and the host cytokine networks. Many viruses, including the human immunodeficiency virus type-1 (HIV-1), rely on the liberation and cellular action of host immune cytokines to expand their host cell range, to regulate their cellular expression, and to maintain their dormant state until the proper extracellular conditions arise. As again exemplified by HIV-1, viruses may also take an active role regulating cytokine expression and cell surface cytokine receptors. Because the viral life cycle, and in particular the HIV-1 life cycle, is so intertwined with cytokine regulatory networks, these networks represent potential points for therapeutic intervention. As our understanding of cellular cytokine pathways involved in viral infection and replication continues to expand, so too will our ability to design rational anti-viral therapies to alter multiple steps along the viral life cycle.     

Bacterial entry into cells: a role for the endocytic machinery

Increasing evidence indicates that pathogens have evolved highly efficient strategies to induce their internalization within host cells. Viruses and bacteria express and expose on their surface, molecules that mimic endogenous ligands to cell receptors, thereby inducing specific intracellular signalling cascades. More recently it has become clear that, as most viruses, bacteria can enter cells via the clathrin-mediated pathway, indicating a key role for endocytosis in pathogens entry into cells. Here we review the pathways followed by Listeria monocytogenes to enter into non-phagocytic cells, as a model for the subversion of cellular functions to induce pathogens internalization.     

DNA sensor cGAS-mediated immune recognition

The host takes use of pattern recognition receptors (PRRs) to defend against pathogen invasion or cellular damage. Among microorganism-associated molecular patterns detected by host PRRs, nucleic acids derived from bacteria or viruses are tightly supervised, providing a fundamental mechanism of host defense. Pathogenic DNAs are supposed to be detected by DNA sensors that induce the activation of NFκB or TBK1-IRF3 pathway. DNA sensor cGAS is widely expressed in innate immune cells and is a key sensor of invading DNAs in several cell types. cGAS binds to DNA, followed by a conformational change that allows the synthesis of cyclic guanosine monophosphate–adenosine monophosphate (cGAMP) from adenosine triphosphate and guanosine triphosphate. cGAMP is a strong activator of STING that can activate IRF3 and subsequent type I interferon production. Here we describe recent progresses in DNA sensors especially cGAS in the innate immune responses against pathogenic DNAs.     

病毒功能性受体的筛选新策略及其应用

病毒通过与靶细胞表面的特异性受体结合,进而吸附、入侵靶细胞,劫持细胞的复制机器完成自身的复制周期。细胞受体作为病毒入侵宿主的门户,细胞受体的结构与功能及其介导病毒入侵的机制一直是病毒学研究的热点之一。对细胞受体的研究有助于了解病毒的致病机制并为病毒性疾病的防控提供科学依据。近年来,利用功能缺失性基因组学筛选病毒功能性受体的进展极大地拓展了人类对病毒入侵机制的认识和理解。目前,应用较为广泛的病毒功能性受体筛选策略包括RNA干扰技术、利用单倍体细胞系随机插入逆转录病毒致突变技术以及基于CRISPR/Cas9系统的新型编辑技术。本文系统介绍并比较了这三种病毒功能性受体筛选新策略,同时对其应用及优缺点进行了归纳总结,可为相关研究者提供一定的参考。     

Viral immunoreceptor tyrosine-based activation motif (ITAM)-mediated signaling in cell transformation and cancer

Viruses frequently co-opt host cell pathways to enhance their propagation or to enable latent infection. Certain receptors expressed by hematopoietic cells have immunoreceptor tyrosine-based activation motifs (ITAMs) in their cytoplasmic domains that initiate cellular activation, proliferation and differentiation. Some viruses have evolved, or acquired from their host, genes that encode ITAM-bearing proteins. These ITAM-bearing viral proteins have been implicated in cellular transformation in virus-infected hematopoietic cells, typically B cells, but also in non-hematopoietic tissues--including endothelial and epithelial cells.     

Novel Insights into Cell Entry of Emerging Human Pathogenic Arenaviruses

Viral hemorrhagic fevers caused by emerging RNA viruses of the Arenavirus family are among the most devastating human diseases. Climate change, global trade, and increasing urbanization promote the emergence and re-emergence of these human pathogenic viruses. Emerging pathogenic arenaviruses are of zoonotic origin and reservoir-to-human transmission is crucial for spillover into human populations. Host cell attachment and entry are the first and most fundamental steps of every virus infection and represent major barriers for zoonotic transmission. During host cell invasion, viruses critically depend on cellular factors, including receptors, co-receptors, and regulatory proteins of endocytosis. An in-depth understanding of the complex interaction of a virus with cellular factors implicated in host cell entry is therefore crucial to predict the risk of zoonotic transmission, define the tissue tropism, and assess disease potential. Over the past years, investigation of the molecular and cellular mechanisms underlying host cell invasion of human pathogenic arenaviruses uncovered remarkable viral strategies and provided novel insights into viral adaptation and virus–host co-evolution that will be covered in the present review.     

Successful control of viruses by NK cells--a balance of opposing forces?

Natural killer (NK) cells play a crucial role in limiting the severity of diseases caused by a range of viruses. Recent data have shown that the effector functions of NK cells can be specifically stimulated when NK cell activation receptors engage cellular major histocompatibility complex (MHC) class I-like ligands induced after infection or by specific viral gene products. However, to counter this NK cell response viruses have evolved an array of strategies to subvert efficient NK cell activation. These data indicate that the balance of host NK cell responses and viral NK cell escape mechanisms can be strategically poised as each strives for survival.     

Sialic acids: Key determinants for invasion by the Apicomplexa

Sialic acids are ubiquitously found on the surface of all vertebrate cells at the extremities of glycan chains and widely exploited by viruses and bacteria to enter host cells. Carbohydrate-bearing receptors are equally important for host cell invasion by the obligate intracellular protozoan parasites of the phylum Apicomplexa. Host cell entry is an active process relying crucially on proteins that engage with receptors on the host cell surface and promote adhesion and internalisation. Assembly into complexes, proteolytic processing and oligomerization are important requirements for the functionality of these adhesins. The combination of adhesive proteins with varying stringency in specificity confers some flexibility to the parasite in face of receptor heterogeneity and immune pressure. Sialic acids are now recognised to critically contribute to selective host cell recognition by various species of the phylum.     

Utilization of human DC-SIGN and L-SIGN for entry and infection of host cells by the New World arenavirus,Junín virus

The target cell tropism of enveloped viruses is regulated by interactions between viral proteins and cellular receptors determining susceptibility at a host cell, tissue or species level. However, a number of additional cell-surface moieties can also bind viral envelope glycoproteins and could act as capture receptors, serving as attachment factors to concentrate virus particles on the cell surface, or to disseminate the virus infection to target organs or susceptible cells within the host. Here, we used Junín virus (JUNV) or JUNV glycoprotein complex (GPC)-pseudotyped particles to study their ability to be internalized by the human C-type lectins hDC- or hL-SIGN. Our results provide evidence that hDC- and hL-SIGN can mediate the entry of Junín virus into cells, and may play an important role in virus infection and dissemination in the host.     

Co-ordinate action of bacterial adhesins and human carcinoembryonic antigen receptors in enhanced cellular invasion by capsulate serum resistant Neisseria meningitidis

Neisseria meningitidis (Nm) is a human specific opportunistic pathogen that occasionally penetrates mucosal barriers via the action of adhesins and invasins and evades host immune mechanisms during further dissemination via capsule expression. From in vitro studies, the primary adhesion of capsulate bacteria is believed to be mediated by polymeric pili, followed by invasion via outer membrane adhesins such as Opa proteins. As the latter requires the surface capsule to be down-modulated, invading bacteria would be serum sensitive and thus avirulent. However, there is recent evidence that capsulate bacteria may interact via Opa proteins when host cells express high levels of carcinoembryonic antigen-related cell adhesion molecules (CEACAMs), their target receptors. Such a situation may arise following increased circulation of inflammatory cytokines that upregulate certain adhesion molecules on host cells. In this study, using a tetracycline controlled expression system, we have developed cell lines with inducible CEACAM expression to mimic post-inflammation state of target tissues and analysed the interplay between the three surface components capsule, pili and Opa proteins in cellular interactions. With two distinct cell lines, not only the level but also the rate of adhesion of capsulate Opa-expressing Nm increased concurrently with CEACAM density. Moreover, when threshold levels of receptor were reached, cellular invasion ensued in an Opa-dependent manner. In studies with cell lines intrinsically expressing pilus receptors, notable synergism in cellular interactions between pili and Opa of several meningococcal strains was observed and was independent of capsule type. A number of internalized bacteria were shown to express capsule and when directly isolated from host cells, these bacteria were as serum resistant as the inoculated phenotype. Furthermore, we observed that agents that block Opa-CEACAM binding substantially reduced cellular invasion, while maintaining a low level of cellular adhesion. These studies highlight some of the factors that may determine increased host susceptibility to infection by serum resistant phenotypes; and demonstrate the potential of selective inhibition of key interactions in preventing target tissue penetration while maintaining a level of colonization.     

Invasive and adherent bacterial pathogens co-Opt host clathrin for infection

Infection by the bacterium Listeria monocytogenes depends on host cell clathrin. To determine whether this requirement is widespread, we analyzed infection models using diverse bacteria. We demonstrated that bacteria that enter cells following binding to cellular receptors (termed "zippering" bacteria) invade in a clathrin-dependent manner. In contrast, bacteria that inject effector proteins into host cells in order to gain entry (termed "triggering" bacteria) invade in a clathrin-independent manner. Strikingly, enteropathogenic Escherichia coli (EPEC) required clathrin to form actin-rich pedestals in host cells beneath adhering bacteria, even though this pathogen remains extracellular. Furthermore, clathrin accumulation preceded the actin rearrangements necessary for Listeria entry. These data provide evidence for a clathrin-based entry pathway allowing internalization of large objects (bacteria and ligand-coated beads) and used by "zippering" bacteria as part of a general mechanism to invade host mammalian cells. We also revealed a nonendocytic role for clathrin required for extracellular EPEC infections.     

Efficient Infection Mediated by Viral Receptors Incorporated into Retroviral Particles

Many host cell surface proteins, including viral receptors, are incorporated into enveloped viruses. To address the functional significance of these host proteins, murine leukemia viruses containing the cellular receptors for Rous sarcoma virus (Tva) or ecotropic murine leukemia virus (MCAT-1) were produced. These receptor-pseudotyped viruses efficiently infect cells expressing the cognate viral envelope glycoproteins, with titers of up to 10⁵ infectious units per milliliter for the Tva pseudotypes. Receptor and viral glycoprotein specificity and functional requirements are maintained, suggesting that receptor pseudotype infection recapitulates events of normal viral entry. The ability of the Tva and MCAT-1 pseudotypes to infect cells efficiently suggests that, in contrast to human immunodeficiency virus type 1 entry, neither of these retroviral receptors requires a coreceptor for membrane fusion. In addition, the ability of receptor pseudotypes to target infected cells suggests that they may be useful therapeutic reagents for directing infection of viral vectors. Receptor-pseudotyped viruses may be useful for identifying new viral receptors or for defining functional requirements of known receptors. Moreover, this work suggests that the production of receptor pseudotypes in vivo could provide a mechanism for expanded viral tropism with potential effects on the pathogenesis and evolution of the virus.     

Molecular anatomy of mouse hepatitis virus persistence: coevolution of increased host cell resistance and virus virulence.

Persistent infection of murine astrocytoma (DBT) cells with mouse hepatitis virus (MHV) has been established. From this in vitro virus-host system, persistence is mediated at the level of cellular MHV receptor (MHVR) expression and increased virus virulence. MHV persistence selects for resistant host cell populations which abate virus replication. Reductions in MHVR expression were significantly associated with increased host resistance, and transfection of MHVR into resistant host cells completely restored the capacity of cells to support efficient replication of MHV strain A59. The emergence of resistant host cells coselected for variant viruses that had increased avidity for MHVR and also recognized different receptors for entry into resistant cells. These data illustrate that MHV persistence in vitro provides a model to identify critical sites of virus-host interaction at the cellular level which are altered during the evolution of host cell resistance to viral infection and the coevolution of virus virulence.     

Biochemical engineering of the N-acyl side chain of sialic acid: biological implications

N-Acetylneuraminic acid is the most prominent sialic acid in eukaryotes. The structural diversity of sialic acid is exploited by viruses, bacteria, and toxins and by the sialoglycoproteins and sialoglycolipids involved in cell-cell recognition in their highly specific recognition and binding to cellular receptors. The physiological precursor of all sialic acids is N-acetyl D-mannosamine (ManNAc). By recent findings it could be shown that synthetic N-acyl-modified D-mannosamines can be taken up by cells and efficiently metabolized to the respective N-acyl-modified neuraminic acids in vitro and in vivo. Successfully employed D-mannosamines with modified N-acyl side chains include N-propanoyl- (ManNProp), N-butanoyl- (ManNBut)-, N-pentanoyl- (ManNPent), N-hexanoyl- (ManNHex), N-crotonoyl- (ManNCrot), N-levulinoyl- (ManNLev), N-glycolyl- (ManNGc), and N-azidoacetyl D-mannosamine (ManNAc-azido). All of these compounds are metabolized by the promiscuous sialic acid biosynthetic pathway and are incorporated into cell surface sialoglycoconjugates replacing in a cell type-specific manner 10-85% of normal sialic acids. Application of these compounds to different biological systems has revealed important and unexpected functions of the N-acyl side chain of sialic acids, including its crucial role for the interaction of different viruses with their sialylated host cell receptors. Also, treatment with ManNProp, which contains only one additional methylene group compared to the physiological precursor ManNAc, induced proliferation of astrocytes, microglia, and peripheral T-lymphocytes. Unique, chemically reactive ketone and azido groups can be introduced biosynthetically into cell surface sialoglycans using N-acyl-modified sialic acid precursors, a process offering a variety of applications including the generation of artificial cellular receptors for viral gene delivery. This group of novel sialic acid precursors enabled studies on sialic acid modifications on the surface of living cells and has improved our understanding of carbohydrate receptors in their native environment. The biochemical engineering of the side chain of sialic acid offers new tools to study its biological relevance and to exploit it as a tag for therapeutic and diagnostic applications.     

PRRs in pathogen recognition

The innate immune system provides the first line of host defense against invading microorganisms before the development of adaptive immune responses. Innate immune responses are initiated by germline-encoded pattern recognition receptors (PRRs), which recognize specific structures of microorganisms. Toll-like receptors (TLRs) are pattern-recognition receptors that sense a wide range of microorganisms, including bacteria, fungi, protozoa and viruses. TLRs exist either on the cell surface or in the lysosome/endosome compartment and induce innate immune responses. Recently, cytoplasmic PRRs have been identified which detect pathogens that have invaded the cytosol. This review focuses on the pathogen recognition of PRRs in innate immunity.