Biology subjects

Kaufer, B.

Publications and source records attributed to Kaufer, B..

6 recordsLinked to original sources

bioRxiv2026

Opposing roles of ACE2 and TMPRSS2 in IBV Beaudette infection

Infectious bronchitis virus (IBV) is an economically important avian gammacoronavirus. IBV uses sialic acids and heparan sulfate for cellular attachment, but the host proteins governing its infection remain poorly defined. Here, we investigated the roles of angiotensin-converting enzyme 2 (ACE2) and transmembrane serine protease 2 (TMPRSS2) during IBV Beaudette (IBV BD) infection. ACE2 knockout impaired IBV replication in DF-1 cells, whereas ACE2 reconstitution restored infection. Furthermore, ACE2 overexpression enhanced infection of DF-1 and primary chicken embryo kidney cells, supporting the role of chicken ACE2 as a proviral host factor. In contrast, TMPRSS2 exerted a restrictive function on IBV BD infection as demonstrated in TMPRSS2 KO DF-1 cells and upon pharmacological inhibition, which significantly increased IBV BD infection in DF-1 and Vero cells. RNA sequencing revealed significant changes in lipid metabolism and membrane-related processes following TMPRSS2 knockout and pharmacological inhibition. These findings highlight the opposing roles of ACE2 and TMPRSS2 in IBV BD infection, emphasizing the combined impact of proviral host factors, protease activity, and the cellular membrane environment during gammacoronavirus infections.

microbiology↗

bioRxiv2024

Repression of varicella zoster virus gene expression during quiescent infection in the absence of detectable histone deposition

Varicella zoster virus (VZV) is a human-specific herpesvirus that establishes latency in peripheral neurons. The only transcripts detected in infected human trigeminal ganglia (TG) obtained shortly after death correspond to the VZV latency-associated transcript (VLT) and associated VLT-ORF63 splice variants. In vitro studies showed that VLT-ORF63 is translated into a protein (pVLT-ORF63) that induces VZV transcription. The mechanisms that lead to this restricted gene expression and the transition to lytic replication remain unknown, partly due to the difficulty of working with human neurons. In this study, we addressed whether the neuroblastoma-derived cell line SH-SY5Y could serve as a model to investigate the mechanisms that lead to repression of VZV gene expression followed by reactivation. VZV productively infected differentiated SH-SY5Y (dSH-SY5Y) whereas incubation with acyclovir (ACV) inhibited virus replication and induced a progressive repression of the virus. Upon removal of ACV there was production of viral particles in a subset of cells, while others contained non-replicating VZV genomes and VLT-containing transcripts for at least 20 days post-infection (dpi). Exogenous expression of VLT-ORF63 induced productive infection, suggesting that the non-replicating and repressed genomes remained functional. Interestingly, histone deposition was undetectable at VZV genomes in quiescently infected dSH-SY5Y cells, pointing to a potential novel mechanism leading to VZV repression in this neuronal setting.

microbiology↗

bioRxiv2023

First virological and pathological study of Göttingen Minipigs with Dippity Pig Syndrome (DPS)

Dippity Pig Syndrome (DPS) is a well-known but rare complex of clinical signs affecting minipigs, which has not been thoroughly investigated yet. Clinically affected animals show acute appearance of red, exudating lesions across the spine. The lesions are painful, evidenced by arching of the back (dipping), and the onset of clinical symptoms is generally sudden. In order to understand the pathogenesis, histological and virological investigations were performed in affected and unaffected Gottingen Minipigs (GoMPs). The following DNA viruses were screened for using PCR-based methods: Porcine cytomegalovirus (PCMV), which is a porcine roseolovirus (PCMV/PRV), porcine lymphotropic herpesviruses (PLHV-1, PLHV-2, PLHV-3), porcine circoviruses (PCV1, PCV2, PCV3, PCV4), porcine parvovirus 1 (PPV1), and Torque Teno sus virus (TTSuV1, TTSuV2). Screening was also performed for integrated porcine endogenous retroviruses (PERV-A, PERV-B, PERV-C) and recombinant PERV-A/C and their expression as well as for the RNA viruses hepatitis E virus (HEV) and SARS-CoV-2. Eight clinically affected and one unaffected GoMPs were analyzed. Additional unaffected minipigs had been analyzed in the past. The analyzed GoMPs contained PERV-A and PERV-B integrated in the genome, which are present in all pigs and PERV-C, which is present in most, but not all pigs. In one affected GoMPs recombinant PERV-A/C was detected in blood. In this animal a very high expression of PERV mRNA was observed. PCMV/PRV was found in three affected animals, PCV1 was found in three animals with DPS and in the healthy minipig, and PCV3 was detected in two animals with DPS and in the unaffected minipig. Most importantly, in one animal only PLHV-3 was detected. It was found in the affected and unaffected skin, and in other organs. Unfortunately, PLHV-3 could not be studied in all other affected minipigs. None of the other viruses were detected and using electron microscopy, no virus particles were found in the affected skin. This data identified some virus infections in GoMPs with DPS and assign a special role to PLHV-3. Since PCMV/PRV, PCV1, PCV3 and PLHV-3 were also found in unaffected animals, a multifactorial cause of DPS is suggested. However, elimination of the viruses from GoMPs may prevent DPS.

microbiology↗

bioRxiv2020

Left-handed DNA-PAINT for improved superresolution imaging in the nucleus.

DNA point accumulation in nanoscale topography (DNA-PAINT) advances super-resolution microscopy with superior resolution and multiplexing capabilities. However, cellular DNA may interfere with this single-molecule localization technique based on DNA-DNA hybridization. Here, we introduce left-handed DNA (L-DNA) oligomers that do not hybridize to naturally present R-DNA and demonstrate that L-DNA PAINT has the same specificity and multiplexing capability as R-DNA PAINT, but greatly improves specific visualization of nuclear target molecules.

biophysics↗

bioRxiv2018

Role of PML-Nuclear Bodies in Human Herpesvirus 6A and 6B Genome Integration.

Human herpesviruses 6A and 6B (HHV-6A/B) are two betaherpesviruses that readily integrate their genomes into the telomeres of human chromosomes. To date, the cellular or viral proteins that facilitate HHV-6A/B integration remain elusive. In the present study, we demonstrate that the immediate early protein 1 (IE1) of HHV-6A/B colocalizes with telomeres during infection. Moreover, IE1 associates with PML-NBs, a nuclear complex that regulates multiples cellular mechanism including DNA repair and antiviral responses. Furthermore, we could demonstrate that IE1 targets all PML isoforms and that both proteins colocalize at telomeres. To determine the role of PML in HHV-6A/B integration, we generated PML knockout cell lines using CRISPR/Cas9. Intriguingly, in the absence of PML, the IE1 protein could still localize to telomeres albeit less frequently. More importantly, HHV-6A/B integration was impaired in the absence of PML, indicating that it plays a role in the integration process. Taken together, we identified the first cellular protein that aids in the integration of HHV-6A/B and shed light on this targeted integration mechanism.\n\nAuthor summaryHuman herpesviruses type 6A and 6B are relatively common viruses whose infections can be life threatening in patients with a compromised immune system. A rather unique feature of these viruses is their ability to integrate their genome in human chromosomes. Integration takes place is a specialized region of the chromosomes known as telomeres, a region that controls cellular lifespan. To date, the mechanisms leading to HHV-6A and HHV-6B integration remain elusive. Our laboratory has identified that the IE1 protein of HHV-6A and HHV-6B target the telomeres. Moreover, we have shown that IE1 associates with a cellular protein, PML, that is responsible for the regulation of important cellular mechanisms such as the life span of cells and DNA repair. Hence, we studied the role of PML in HHV-6 integration. Our study demonstrates that in absence of PML, the HHV-6A and HHV-6B integrate 50-70% less frequently. Thus, our study unveils the first cellular protein involved in HHV-6A and HHV-6 chromosomal integration.

microbiology↗

bioRxiv2018

Targeted induction of de novo Fatty acid synthesis enhances MDV replication in a COX-2/PGE2 dependent mechanism through EP2 and EP4 receptors engagement

Many viruses alter de novo Fatty Acid (FA) synthesis pathway, which can increase availability of energy for replication and provide specific cellular substrates for particle assembly. Mareks disease virus (MDV) is a herpesvirus that causes deadly lymphoma and has been linked to alterations of lipid metabolism in MDV-infected chickens. However, the role of lipid metabolism in MDV replication is largely unknown. We demonstrate here that infection of primary chicken embryonic fibroblast with MDV activates de novo lipogenesis, which is required for virus replication. In contrast, activation of Fatty Acid Oxidation (FAO) reduced MDV titer, while inhibition of FAO moderately increased virus replication. Thus optimized virus replication occurs if synthetized fatty acids are not used for generation of energy in the infected cells, and they are likely converted to lipid compounds, which are important for virus replication. We showed that infection with MDV activates COX-2/PGE2 pathway and increases the biosynthesis of PGE2, a lipid mediator generated from arachidonic acid. Inhibition of COX-2 or PGE2 receptors, namely EP2 and EP4 receptors, reduced MDV titer, indicating that COX-2/PGE2 pathway are involved in virus replication. Our data show that the FA synthesis pathway inhibitors reduce COX-2 expression level and PGE2 synthesis in MDV infected cells, arguing that there is a direct link between virus-induced fatty acid synthesis and activation of COX-2/PGE2 pathway. This notion was confirmed by the results showing that PGE2 can restore MDV replication in the presence of the FA synthesis pathway inhibitors. Taken together, our data demonstrate that MDV uses FA synthesis pathway to enhance PGE2 synthesis and promote MDV replication through EP2 and EP4 receptors engagement.

microbiology↗