A Genome-Wide Haploid Genetic Screen Identifies Heparan Sulfate-Associated Genes and the Macropinocytosis Modulator TMED10 as Factors Supporting Vaccinia Virus Infection.

Vaccinia virus is a promising viral vaccine and gene delivery candidate and has historically been used as a model to study poxvirus-host cell interactions. We employed a genome-wide insertional mutagenesis approach in human haploid cells to identify host factors crucial for vaccinia virus infection. A library of mutagenized HAP1 cells was exposed to modified vaccinia virus Ankara (MVA).

A combinational CRISPR/Cas9 gene-editing approach can halt HIV replication and prevent viral escape.

HIV presents one of the highest evolutionary rates ever detected and combination antiretroviral therapy is needed to overcome the plasticity of the virus population and control viral replication. Conventional treatments lack the ability to clear the latent reservoir, which remains the major obstacle towards a cure. Novel strategies, such as CRISPR/Cas9 gRNA-based genome-editing, can permanently disrupt the HIV genome.

The E3 Ubiquitin Ligase TMEM129 Is a Tri-Spanning Transmembrane Protein.

Misfolded proteins from the endoplasmic reticulum (ER) are transported back into the cytosol for degradation via the ubiquitin-proteasome system. The human cytomegalovirus protein US11 hijacks this ER-associated protein degradation (ERAD) pathway to downregulate human leukocyte antigen (HLA) class I molecules in virus-infected cells, thereby evading elimination by cytotoxic T-lymphocytes. Recently, we identified the E3 ubiquitin ligase transmembrane protein 129 (TMEM129) as a key player in this process, where interference with TMEM129 activity in human cells completely abrogates US11-mediated class I degradation.

Comprehensive profiling of functional Epstein-Barr virus miRNA expression in human cell lines.

Background: Epstein-Barr virus (EBV) establishes lifelong infections in its human host. The virus is associated with a broad range of malignancies of lymphoid and epithelial origin, including Burkitt's lymphoma, post-transplant lymphoproliferative disease, nasopharyngeal carcinoma and gastric carcinoma. During the latent phase of its life cycle, EBV expresses more than 40 mature miRNAs that are highly abundant in tumor cells and may contribute to oncogenesis.

CRISPR/Cas9-Mediated Genome Editing of Herpesviruses Limits Productive and Latent Infections.

Herpesviruses infect the majority of the human population and can cause significant morbidity and mortality. Herpes simplex virus (HSV) type 1 causes cold sores and herpes simplex keratitis, whereas HSV-2 is responsible for genital herpes. Human cytomegalovirus (HCMV) is the most common viral cause of congenital defects and is responsible for serious disease in immuno-compromised individuals.

The Epstein-Barr Virus Glycoprotein gp150 Forms an Immune-Evasive Glycan Shield at the Surface of Infected Cells.

Cell-mediated immunity plays a key role in host control of viral infection. This is exemplified by life-threatening reactivations of e.g.

Recent advances in viral evasion of the MHC Class I processing pathway.

T-cell mediated adaptive immunity against viruses relies on recognition of virus-derived peptides by CD4(+) and CD8(+) T cells. Detection of pathogen-derived peptide-MHC-I complexes triggers CD8(+) T cells to eliminate the infected cells. Viruses have evolved several mechanisms to avoid recognition, many of which target the MHC-I antigen-processing pathway.

Immune Evasion by Epstein-Barr Virus.

Epstein-Bar virus (EBV) is widespread within the human population with over 90% of adults being infected. In response to primary EBV infection, the host mounts an antiviral immune response comprising both innate and adaptive effector functions. Although the immune system can control EBV infection to a large extent, the virus is not cleared.

Suppression of a Natural Killer Cell Response by Simian Immunodeficiency Virus Peptides.

Natural killer (NK) cell responses in primates are regulated in part through interactions between two highly polymorphic molecules, the killer-cell immunoglobulin-like receptors (KIRs) on NK cells and their major histocompatibility complex (MHC) class I ligands on target cells. We previously reported that the binding of a common MHC class I molecule in the rhesus macaque, Mamu-A1*002, to the inhibitory receptor Mamu-KIR3DL05 is stabilized by certain simian immunodeficiency virus (SIV) peptides, but not by others. Here we investigated the functional implications of these interactions by testing SIV peptides bound by Mamu-A1*002 for the ability to modulate Mamu-KIR3DL05+ NK cell responses.

Pathogenicity of Bovine Neonatal Pancytopenia-associated vaccine-induced alloantibodies correlates with Major Histocompatibility Complex class I expression.

Bovine Neonatal Pancytopenia (BNP), a fatal bleeding syndrome of neonatal calves, is caused by maternal alloantibodies absorbed from colostrum and is characterized by lymphocytopenia, thrombocytopenia and bone marrow hypoplasia. An inactivated viral vaccine is the likely source of alloantigens inducing BNP-associated alloantibodies in the dam. In this study the specificity of BNP alloantibodies was assessed and was linked to the pathology of BNP.

Proteasomal Degradation of Proinsulin Requires Derlin-2, HRD1 and p97.

Patients with type 1 diabetes (T1D) suffer from beta-cell destruction by CD8+ T-cells that have preproinsulin as an important target autoantigen. It is of great importance to understand the molecular mechanism underlying the processing of preproinsulin into these CD8+ T-cell epitopes. We therefore studied a pathway that may contribute to the production of these antigenic peptides: degradation of proinsulin via ER associated protein degradation (ERAD).

Viral immune evasion: Lessons in MHC class I antigen presentation.

The MHC class I antigen presentation pathway enables cells infected with intracellular pathogens to signal the presence of the invader to the immune system. Cytotoxic T lymphocytes are able to eliminate the infected cells through recognition of pathogen-derived peptides presented by MHC class I molecules at the cell surface. In the course of evolution, many viruses have acquired inhibitors that target essential stages of the MHC class I antigen presentation pathway.

Viral inhibition of the transporter associated with antigen processing (TAP): a striking example of functional convergent evolution.

Herpesviruses are large DNA viruses that are highly abundant within their host populations. Even in the presence of a healthy immune system, these viruses manage to cause lifelong infections. This persistence is partially mediated by the virus entering latency, a phase of infection characterized by limited viral protein expression.

Silencing the shutoff protein of Epstein-Barr virus in productively infected B cells points to (innate) targets for immune evasion.

During productive infection with Epstein-Barr virus (EBV), a dramatic suppression of cellular protein expression is caused by the viral alkaline exonuclease BGLF5. Among the proteins downregulated by BGLF5 are multiple immune components. Here, we show that shutoff reduces expression of the innate EBV-sensing Toll-like receptor-2 and the lipid antigen-presenting CD1d molecule, thereby identifying these proteins as novel targets of BGLF5.

Cowpox virus protein CPXV012 eludes CTLs by blocking ATP binding to TAP.

CD8(+) CTLs detect virus-infected cells through recognition of virus-derived peptides presented at the cell surface by MHC class I molecules. The cowpox virus protein CPXV012 deprives the endoplasmic reticulum (ER) lumen of peptides for loading onto newly synthesized MHC class I molecules by inhibiting the transporter associated with Ag processing (TAP). This evasion strategy allows the virus to avoid detection by the immune system.

A high-coverage shRNA screen identifies TMEM129 as an E3 ligase involved in ER-associated protein degradation.

Misfolded ER proteins are retrotranslocated into the cytosol for degradation via the ubiquitin-proteasome system. The human cytomegalovirus protein US11 exploits this ER-associated protein degradation (ERAD) pathway to downregulate HLA class I molecules in virus-infected cells, thereby evading elimination by cytotoxic T-lymphocytes. US11-mediated degradation of HLA class I has been instrumental in the identification of key components of mammalian ERAD, including Derlin-1, p97, VIMP and SEL1L.

Epstein-Barr virus large tegument protein BPLF1 contributes to innate immune evasion through interference with toll-like receptor signaling.

Viral infection triggers an early host response through activation of pattern recognition receptors, including Toll-like receptors (TLR). TLR signaling cascades induce production of type I interferons and proinflammatory cytokines involved in establishing an anti-viral state as well as in orchestrating ensuing adaptive immunity. To allow infection, replication, and persistence, (herpes)viruses employ ingenious strategies to evade host immunity.

TAP mediates import of Mycobacterium tuberculosis-derived peptides into phagosomes and facilitates loading onto HLA-I.

Processing and presentation of antigen on MHC-I class I molecules serves to present peptides derived from cytosolic proteins to CD8(+) T cells. Infection with bacteria that remain in phagosomal compartments, such as Mycobacterium tuberculosis (Mtb), provides a challenge to this immune recognition as bacterial proteins are segregated from the cytosol. Previously we identified the Mtb phagosome itself as an organelle capable of loading MHC Class I molecules with Mtb antigens.

MHC class I molecules are preferentially ubiquitinated on endoplasmic reticulum luminal residues during HRD1 ubiquitin E3 ligase-mediated dislocation.

Misfolded MHC class I heavy chains (MHC I HCs) are targeted for endoplasmic reticulum (ER)-associated degradation (ERAD) by the ubiquitin E3 ligase HRD1, and E2 ubiquitin conjugating enzyme UBE2J1, and represent one of the few known endogenous ERAD substrates. The mechanism by which misfolded proteins are dislocated across the ER membrane into the cytosol is unclear. Here, we investigate the requirements for MHC I ubiquitination and degradation and show that endogenous misfolded MHC I HCs are recognized in the ER lumen by EDEM1 in a glycan-dependent manner and targeted to the core SEL1L/HRD1/UBE2J1 complex.

The comparative genomics of human respiratory syncytial virus subgroups A and B: genetic variability and molecular evolutionary dynamics.

Genomic variation and related evolutionary dynamics of human respiratory syncytial virus (RSV), a common causative agent of severe lower respiratory tract infections, may affect its transmission behavior. RSV evolutionary patterns are likely to be influenced by a precarious interplay between selection favoring variants with higher replicative fitness and variants that evade host immune responses. Studying RSV genetic variation can reveal both the genes and the individual codons within these genes that are most crucial for RSV survival.

Genetically engineered human islets protected from CD8-mediated autoimmune destruction in vivo.

Islet transplantation is a promising therapy for type 1 diabetes, but graft function and survival are compromised by recurrent islet autoimmunity. Immunoprotection of islets will be required to improve clinical outcome. We engineered human β cells to express herpesvirus-encoded immune-evasion proteins, "immunevasins.

Viral interference with antigen presentation: trapping TAP.

Following primary infection, herpesviruses persist for life in their hosts, even when vigorous anti-viral immunity has been induced. Failure of the host immune system to eliminate infected cells is facilitated by highly effective immune evasion strategies acquired by these herpesviruses during millions of years of co-evolution with their hosts. Here, we review the mechanisms of action of viral gene products that lead to cytotoxic T cell evasion through interference with the function of the transporter associated with antigen processing, TAP.