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Holoubek, J.

Publications and source records attributed to Holoubek, J..

3 recordsLinked to original sources

The influence of the pre-membrane and envelope proteins on structure, pathogenicity and tropism of tick-borne encephalitis virus

Tick-borne encephalitis virus (TBEV) is a neurotropic flavivirus that causes thousands of human infections annually. Viral tropism in the brain is determined by the presence of necessary receptors, entry factors and the ability of the virus to overcome host defenses. The viral structural proteins, pre-membrane (prM) and envelope (E), play an important role in receptor binding, membrane fusion, particle maturation, and antibody neutralization. To understand how these proteins influence virus distribution and tropism in the brain, we generated a chimeric virus harboring the prM and ectodomain of E from TBEV in the background of the low pathogenic Langat virus (LGTV). We solved the atomic structures of both the chimeric virus and LGTV to compare them to the known TBEV structure. We show that this chimeric virus remains low-pathogenic, while being structurally and antigenically similar to TBEV. Using 3D optical whole brain imaging combined with immunohistochemistry, we found that both LGTV and the chimeric virus primarily infect cerebral cortex, with no significant differences in their localization or tropism. In contrast, TBEV shows high infection of the cerebellum and strong preference towards Purkinje cells, indicating that the non-structural proteins are important for determining TBEV tropism in the brain. Together, this provides new insights into the roles of the structural and non-structural proteins of tick-borne flaviviruses.

microbiology↗

Exploring the Transcriptomic Profile of Human Monkeypox Virus via CAGE and Native RNA Sequencing Approaches

In this study, we employed short- and long-read sequencing technologies to delineate the transcriptional architecture of the human monkeypox virus and to identify key regulatory elements that govern its gene expression. Specifically, we conducted a transcriptomic analysis to annotate the transcription start sites (TSSs) and transcription end sites (TESs) of the virus by utilizing cap analysis of gene expression sequencing on the Illumina platform and direct RNA sequencing on the Oxford Nanopore technology device. Our investigations uncovered significant complexity in the use of alternative TSSs and TESs in viral genes. In this research, we also detected the promoter elements and poly(A) signals associated with the viral genes. Additionally, we identified novel genes in both the left and right variable regions of the viral genome. ImportanceGenerally, gaining insight into how the transcription of a virus is regulated offers insights into the key mechanisms that control its life cycle. The recent outbreak of the human monkeypox virus has underscored the necessity of understanding the basic biology of its causative agent. Our results are pivotal for constructing a comprehensive transcriptomic atlas of the human monkeypox virus, providing valuable resources for future studies.

microbiology↗

In-depth Temporal Transcriptome Profiling of Monkeypox and Host Cells using Nanopore Sequencing

The recent Monkeypox outbreak showed the importance of studying the basic biology of orthopoxviruses. However, the transcriptome of its causative agent has not been investigated before neither with short-, nor with long-read sequencing approaches. This Oxford Nanopore long-read RNA-Sequencing dataset fills this gap. Our direct cDNA and native RNA sequencing data enable the in-depth characterization of the transcriptomic architecture and dynamics of the gene expressions of monkeypox virus; and also the deeper understanding of the changes it causes in the host cells on a transcriptome level.

microbiology↗