A pioneer of yeast genetics in Croatia: Zoran Zgaga’s contribution to make national research acknowledged worldwide

This study is an attempt to evaluate the pathway and the achievements of yeast genetics in Croatia. The study represents both, an authors’ review and a historical overview and therefore is of value for yeast geneticists aswell as for historians of science

A pioneer of yeast genetics in Croatia: Zoran Zgaga’s contribution to make national research acknowledged worldwide

This study is an attempt to evaluate the pathway and the achievements of yeast genetics in Croatia. The study represents both, an authors’ review and a historical overview and therefore is of value for yeast geneticists aswell as for historians of science

CMV and natural killer cells: shaping the response to vaccination.

Cytomegaloviruses (CMVs) are highly prevalent, persistent human pathogens that not only evade but also shape our immune responses. Natural killer (NK) cells play an important role in the control of CMV and CMVs have in turn developed a plethora of immunoevasion mechanisms targeting NK cells. This complex interplay can leave a long-lasting imprint on the immune system in general and affect responses toward other pathogens and vaccines. This review aims to provide an overview of NK cell biology and development, the manipulation of NK cells by CMVs and the potential impact of these evasion strategies on responses to vaccination

The Cytomegalovirus Tegument Protein UL35 Antagonizes Pattern Recognition Receptor-Mediated Type I IFN Transcription

The rapid activation of pattern recognition receptor (PRR)-mediated type I interferon (IFN) signaling is crucial for the host response to infection. In turn, human cytomegalovirus (HCMV) must evade this potent response to establish life-long infection. Here, we reveal that the HCMV tegument protein UL35 antagonizes the activation of type I IFN transcription downstream of the DNA and RNA sensors cGAS and RIG-I, respectively. We show that ectopic expression of UL35 diminishes the type I IFN response, while infection with a recombinant HCMV lacking UL35 induces an elevated type I IFN response compared to wildtype HCMV. With a series of luciferase reporter assays and the analysis of signaling kinetics upon HCMV infection, we observed that UL35 downmodulates PRR signaling at the level of the key signaling factor TANK-binding kinase 1 (TBK1). Finally, we demonstrate that UL35 and TBK1 co-immunoprecipitate when co-expressed in HEK293T cells. In addition, we show that a previously reported cellular binding partner of UL35, O-GlcNAc transferase (OGT), post-translationally GlcNAcylates UL35, but that this modification is not required for the antagonizing effect of UL35 on PRR signaling. In summary, we have identified UL35 as the first HCMV protein to antagonize the type I IFN response at the level of TBK1, thereby enriching our understanding of how this important herpesvirus escapes host immune responses

Dengue Virus-Infected Dendritic Cells, but Not Monocytes, Activate Natural Killer Cells through a Contact-Dependent Mechanism Involving Adhesion Molecules

Natural killer (NK) cells play a protective role against dengue virus (DENV) infection, but the cellular and molecular mechanisms are not fully understood. Using an optimized humanized mouse model, we show that human NK cells, through the secretion of gamma interferon (IFN-γ), are critical in the early defense against DENV infection. Depletion of NK cells or neutralization of IFN-γ leads to increased viremia and more severe thrombocytopenia and liver damage in humanized mice. In vitro studies using autologous human NK cells show that DENV-infected monocyte-derived dendritic cells (MDDCs), but not monocytes, activate NK cells in a contact-dependent manner, resulting in upregulation of CD69 and CD25 and secretion of IFN-γ. Blocking adhesion molecules (LFA-1, DNAM-1, CD2, and 2β4) on NK cells abolishes NK cell activation, IFN-γ secretion, and the control of DENV replication. NK cells activated by infected MDDCs also inhibit DENV infection in monocytes. These findings show the essential role of human NK cells in protection against acute DENV infection in vivo, identify adhesion molecules and dendritic cells required for NK cell activation, and delineate the sequence of events for NK cell activation and protection against DENV infection.IMPORTANCE Dengue is a mosquito-transmitted viral disease with a range of symptoms, from mild fever to life-threatening dengue hemorrhagic fever. The diverse disease manifestation is thought to result from a complex interplay between viral and host factors. Using mice engrafted with a human immune system, we show that human NK cells inhibit virus infection through secretion of the cytokine gamma interferon and reduce disease pathogenesis, including depletion of platelets and liver damage. During a natural infection, DENV initially infects dendritic cells in the skin. We find that NK cells interact with infected dendritic cells through physical contact mediated by adhesion molecules and become activated before they can control virus infection. These results show a critical role of human NK cells in controlling DENV infection in vivo and reveal the sequence of molecular and cellular events that activate NK cells to control dengue virus infection

Murine CMV Expressing the High Affinity NKG2D Ligand MULT-1: A Model for the Development of Cytomegalovirus-Based Vaccines

The development of a vaccine against human cytomegalovirus (CMV) has been a subject of long-term medical interest. The research during recent years identified CMV as an attractive vaccine vector against infectious diseases and tumors. The immune response to CMV persists over a lifetime and its unique feature is the inflationary T cell response to certain viral epitopes. CMV encodes numerous genes involved in immunoevasion, which are non-essential for virus growth in vitro. The deletion of those genes results in virus attenuation in vivo, which enables us to dramatically manipulate its virulence and the immune response. We have previously shown that the murine CMV (MCMV) expressing RAE-1γ, one of the cellular ligands for the NKG2D receptor, is highly attenuated in vivo but retains the ability to induce a strong CD8+ T cell response. Here, we demonstrate that recombinant MCMV expressing high affinity NKG2D ligand murine UL16 binding protein-like transcript (MULT-1) (MULT-1MCMV) inserted in the place of its viral inhibitor is dramatically attenuated in vivo in a NK cell-dependent manner, both in immunocompetent adult mice and in immunologically immature newborns. MULT-1MCMV was more attenuated than the recombinant virus expressing RAE-1γ. Despite the drastic sensitivity to innate immune control, MULT-1MCMV induced an efficient CD8+ T cell response to viral and vectored antigens. By using in vitro assay, we showed that similar to RAE-1γMCMV, MULT-1 expressing virus provided strong priming of CD8+ T cells. Moreover, MULT-1MCMV was able to induce anti-viral antibodies, which after passing the transplacental barrier protect offspring of immunized mothers from challenge infection. Altogether, this study further supports the concept that CMV expressing NKG2D ligand possesses excellent characteristics to serve as a vaccine or vaccine vector

The herpesviral antagonist m152 reveals differential activation of STING‐dependent IRF and NF‐κB signaling and STING's dual role during MCMV infection

Cytomegaloviruses (CMVs) are master manipulators of the host immune response. Here, we reveal that the murine CMV (MCMV) protein m152 specifically targets the type I interferon (IFN) response by binding to stimulator of interferon genes (STING), thereby delaying its trafficking to the Golgi compartment from where STING initiates type I IFN signaling. Infection with an MCMV lacking m152 induced elevated type I IFN responses and this leads to reduced viral transcript levels both in vitro and in vivo This effect is ameliorated in the absence of STING Interestingly, while m152 inhibits STING-mediated IRF signaling, it did not affect STING-mediated NF-κB signaling. Analysis of how m152 targets STING translocation reveals that STING activates NF-κB signaling already from the ER prior to its trafficking to the Golgi. Strikingly, this response is important to promote early MCMV replication. Our results show that MCMV has evolved a mechanism to specifically antagonize the STING-mediated antiviral IFN response, while preserving its pro-viral NF-κB response, providing an advantage in the establishment of an infection

Cytomegaloviruses Exploit Recycling Rab Proteins in the Sequential Establishment of the Assembly Compartment

Cytomegaloviruses (CMV) reorganize membranous system of the cell in order to develop a virion assembly compartment (VAC). The development starts in the early (E) phase of infection with the reorganization of the endosomal system and the Golgi and proceeds to the late phase until newly formed virions are assembled and released. The events in the E phase involve reorganization of the endosomal recycling compartment (ERC) in a series of cellular alterations that are mostly unknown. In this minireview, we discuss the effect of murine CMV infection on Rab proteins, master regulators of membrane trafficking pathways, which in the cascades with their GEFs and GAPs organize the flow of membranes through the ERC. Immunofluorescence analyzes of murine CMV infected cells suggest perturbations of Rab cascades that operate at the ERC. Analysis of cellular transcriptome in the course of both murine and human CMV infection demonstrates the alteration in expression of cellular genes whose products are known to build Rab cascades. These alterations, however, cannot explain perturbations of the ERC. Cellular proteome data available for human CMV infected cells suggests the potential role of RabGAP downregulation at the end of the E phase. However, the very early onset of the ERC alterations in the course of MCMV infection indicates that CMVs exploit Rab cascades to reorganize the ERC, which represents the earliest step in the sequential establishment of the cVAC

Human Cytomegalovirus Nuclear Capsids Associate with the Core Nuclear Egress Complex and the Viral Protein Kinase pUL97

Abstract: The nuclear phase of herpesvirus replication is regulated through the formation of regulatory multi-component protein complexes. Viral genomic replication is followed by nuclear capsid assembly, DNA encapsidation and nuclear egress. The latter has been studied intensely pointing to the formation of a viral core nuclear egress complex (NEC) that recruits a multimeric assembly of viral and cellular factors for the reorganization of the nuclear envelope. To date, the mechanism of the association of human cytomegalovirus (HCMV) capsids with the NEC, which in turn initiates the specific steps of nuclear capsid budding, remains undefined. Here, we provide electron microscopy-based data demonstrating the association of both nuclear capsids and NEC proteins at nuclear lamina budding sites. Specifically, immunogold labelling of the core NEC constituent pUL53 and NEC-associated viral kinase pUL97 suggested an intranuclear NEC-capsid interaction. Staining patterns with phospho-specific lamin A/C antibodies are compatible with earlier postulates of targeted capsid egress at lamina-depleted areas. Important data were provided by co-immunoprecipitation and in vitro kinase analyses using lysates from HCMV-infected cells, nuclear fractions, or infectious virions. Data strongly suggest that nuclear capsids interact with pUL53 and pUL97. Combined, the findings support a refined concept of HCMV nuclear trafficking and NEC-capsid interaction

The murine cytomegalovirus M35 protein antagonizes type I IFN induction downstream of pattern recognition receptors by targeting NF-κB mediated transcription.

The herpesvirus cytomegalovirus can cause severe morbidity in immunosuppressed people and poses a much greater global problem in the context of congenital infections than the Zika virus. To establish infection, cytomegalovirus needs to modulate the antiviral immune response of its host. One of the first lines of defense against viral infections is the type I interferon response which is activated by cellular sensors called pattern recognition receptors. These receptors sense viral entry and rapidly induce the transcription of type I interferons, which are instrumental for the induction of an antiviral state in infected and surrounding cells. We have identified the first viral protein encoded by murine cytomegalovirus, the M35 protein, that counteracts type I interferon transcription downstream of multiple pattern recognition receptors. We found that this viral countermeasure occurs shortly after viral entry into the host cell, as M35 is delivered with the viral particle. M35 then localizes to the nucleus where it modulates NF-κB-mediated transcription. In vivo, murine cytomegalovirus deficient of the M35 protein replicates to lower levels in spleen and liver and cannot establish a productive infection in the salivary glands, which is a key site of viral transmission, highlighting the important role of M35 for the establishment of infection. Our study provides novel insights into the complex interaction between cytomegalovirus and the innate immune response of its host