Biology subjects

Sutter, S. O.

Publications and source records attributed to Sutter, S. O..

4 recordsLinked to original sources

bioRxiv2025

The critical role of enterovirus 2A protease in viral translation, replication, and antagonism of host antiviral responses.

Enteroviruses dramatically remodel the cellular infrastructure for efficient replication and curtailing host antiviral responses. Proteases 2Apro and 3Cpro have been implicated in these processes based on in vitro studies, ectopic overexpression, and surrogate infection systems, but their relative contributions are unknown. Here, we replace the essential 2A cleavage site at the P1-P2 junction with an internal ribosome entry site (IRES), 3CD cleavage site, or T2A sequence, allowing us to catalytically inactivate 2Apro. Viruses with an inactive 2Apro are hampered in replication in cell lines and are severely attenuated in a Coxsackievirus B3 (CVB3) mouse pancreatitis infection model. We show that 2Apro, but not 3Cpro, is essential during infection for disturbing nucleocytoplasmic transport, shutting down host mRNA translation, suppressing stress granule formation, cleaving retinoic acid-inducible gene-1 (RIG-I)-like receptor (RLR) signaling pathway proteins and suppressing interferon-/{beta} (IFN-/{beta}) transcription, and overcoming the antiviral action of IFN-induced restriction factors. Moreover, using an advanced single-molecule live cell imaging approach, we reveal that 2Apro is important for the initial round of replication of the incoming viral RNA, which is a bottleneck for efficient infection. In conclusion, we establish that 2Apro plays a critical role in subverting antiviral responses and establishing a favorable host environment to expedite enterovirus replication.

microbiology↗

bioRxiv2024

The interferon-γ inducible factor 16 (IFI16) restricts adeno-associated virus serotype 2 (AAV2) transduction in an immune-modulatory independent way

We determined the transcription profile of AAV2-infected primary human fibroblasts. Subsequent analysis revealed that cells respond to AAV infection through changes in several significantly affected pathways including cell cycle regulation, chromatin modulation, and innate immune responses. Various assays were performed to validate selected differentially expressed genes and confirmed not only the quality but also the robustness of the raw data. One of the genes upregulated in AAV2 infected cells was the interferon-{gamma} inducible factor 16 (IFI16). IFI16 is known as a multifunctional cytosolic and nuclear innate immune sensor for double-stranded, as well as single-stranded DNA, exerting its effects through various mechanisms, such as interferon response, epigenetic modifications, or transcriptional regulation. IFI16 thereby constitutes a restriction factor for many different viruses amongst them, as shown here, AAV2 and thereof derived vectors. Indeed, the post-transcriptional silencing of IFI16 significantly increased AAV2 transduction efficiency, independent of the structure of the virus/vector genome. We also show that IFI16 exerts its inhibitory effect on AAV2 transduction in an immune-modulatory independent way, by interfering with Sp1-dependent transactivation of wild-type AAV2 and AAV2 vector promoters. IMPORTANCEAdeno-associated virus (AAV) vectors are among the most frequently used viral vectors for gene therapy. The lack of pathogenicity of the parental virus, the long-term persistence as episomes in non-proliferating cells, and the availability of a variety of AAV serotypes differing in their cellular tropism are advantageous features of this biological nanoparticle. To deepen our understanding of virus-host interactions, especially in terms of innate immune responses, we present here the first transcriptome analysis of AAV serotype 2 (AAV2) infected human primary fibroblasts. Our findings indicate that the interferon-{gamma} inducible factor 16 (IFI16) acts as an antiviral factor in AAV2 infection and AAV2 vector-mediated cell transduction in an immune-modulatory independent way by interrupting the Sp1-dependent gene expression from viral or vector genomes.

microbiology↗

bioRxiv2024

Non-canonical induction of autophagy increases adeno-associated virus type 2 (AAV2) transduction efficiency

Adeno-associated virus (AAV) serotypes infect a wide range of cell types, making this member of the parvovirus family a versatile tool in gene therapy. Infection as well as transduction is set in motion by means of specific receptors in conjunction with trafficking pathways, particularly endocytosis, a main cell entry pathway of non-enveloped viruses. Here, we report that efficacy of transduction is enhanced upon treating cells with hyperosmotic sucrose, a known blocker of clathrin-mediated endocytosis, through the non-canonical induction of autophagy. This mechanism of autophagy induction, however, is different from the previously reported AAV2-mediated induction of autophagy, which relies on a canonical, phosphoinositide 3-kinase class III (PI3K-III) complex-dependent pathway and appears to be dependent on the virus intrinsic secreted phospholipase A2 (sPLA2) domain, particularly its catalytic center activity. IMPORTANCEAdeno-associated virus (AAV) vectors are among the most frequently applied virus-based delivery vehicles for gene therapy. Lack of pathogenicity for humans, availability of a huge number of AAV serotypes differing in their cellular tropism, and the mainly episomal persistence of AAV vector genomes are clear advantages of these biological nanoparticles. By exploring non-pharmacological inducers of autophagy, we provide evidence for a potent and easy to apply strategy to significantly improve the efficacy of recombinant AAV-based gene delivery in hepatic and structural cells. Besides, our data also demonstrate the importance of autophagy for AAV2 infection and vector-mediated transduction in non-hepatic cells.

microbiology↗

bioRxiv2021

Adeno-associated virus type 2 (AAV2) uncoating is a stepwise process and is linked to structural reorganization of the nucleolus

Nucleoli are membrane-less structures located within the nucleus and are known to be involved in many cellular functions, including stress response and cell cycle regulation. Besides, many viruses can employ the nucleolus or nucleolar proteins to promote different steps of their life cycle such as replication, transcription and assembly. While adeno-associated virus type 2 (AAV2) capsids have previously been reported to enter the host cell nucleus and accumulate in the nucleolus, both the role of the nucleolus in AAV2 infection, and the viral uncoating mechanism remain elusive. In all prior studies on AAV uncoating, viral capsids and viral genomes were not directly correlated on the single cell level, at least not in absence of a helper virus. To elucidate the properties of the nucleolus during AAV2 infection and to assess viral uncoating on a single cell level, we combined immunofluorescence analysis for detection of intact AAV2 capsids and capsid proteins with fluorescence in situ hybridization for detection of AAV2 genomes. The results of our experiments provide evidence that uncoating of AAV2 particles occurs in a stepwise process that is completed in the nucleolus and supported by alteration of the nucleolar structure. Author SummaryAdeno-associated virus (AAV) capsids have been reported to enter the host cell nucleus and accumulate in the nucleolus. However, both the role of the nucleolus in AAV2 infection as well as the viral uncoating mechanism remain unknown. Here, we provide evidence that uncoating of the AAV2 particle is a stepwise process that is completed in the nucleolus and supported by alteration of the nucleolar morphology.

molecular biology↗