Biology subjects

Tietz, O.

Publications and source records attributed to Tietz, O..

3 recordsLinked to original sources

bioRxiv2026

Modular PEGylation Confers Reduced Immune-Cell Uptake and Enhanced Pharmacokinetic Performance to Encapsulin Protein Nanocages

Encapsulins are self-assembling prokaryotic protein nanocages with growing potential as systemic drug delivery systems, but their pharmacokinetic behaviour remains poorly characterised, and rapid immune recognition and clearance may limit delivery to target tissues. Here, we show that modular PEGylation of a SpyCatcher-decorated encapsulin Alkaliphilus metalliredigens (Am-S) markedly reduces macrophage uptake and extends systemic circulation. Site-directed surface PEGylation using SpyTagged PEG achieved 82% conjugation efficiency, corresponding to an estimated average of 49 PEG chains per 60 subunit Am-S nanocage, without compromising nanocage assembly, morphology, or colloidal stability. PEGylated Am-S also retained solubility and protein integrity following freeze-thaw cycling and six months of storage. When interacted with RAW 264.7 macrophages in vitro, PEGylation substantially reduced nanocage association and internalisation relative to non-PEGylated nanocages. Following intravenous administration in BALB/c mice, PEGylated nanocages exhibited markedly prolonged circulation, with >50% of the injected dose remaining after 1 h compared with 2.6% for non-PEGylated Am-S, and a circulatory half-life of 1 h 43 min. To our knowledge, this represents the first pharmacokinetic characterisation of an encapsulin nanocage. Together, these findings demonstrate that controlled PEGylation can substantially reduce macrophage interactions and prolong encapsulin circulation in vivo.

bioengineering↗

bioRxiv2025

Importin α/β1 dependent nuclear import of Black Sea Bass Polyomavirus Large Tumor Antigen is mediated by a classical NLS located downstream of the SF3 helicase domain

Polyomaviruses (PyVs) are small dsDNA viruses that replicate in the host cell nucleus, primarily relying on the viral encoded large tumor antigen (LTA). Aided by recent advances in molecular biology techniques, the list of known PyVs is rapidly growing, revealing unexpected broad sequence, and host heterogenicity. Given their dependence on nuclear localization, large tumor antigens represent an attractive model for studying the nuclear transport process. A comprehensive analysis of the evolution of classical nuclear localization signals (cNLSs) within LTAs encoded by PyVs infecting mammals highlighted strong positional conservation of cNLSs between the LXCXE motif and the origin-binding domain (OBD). Here we extend such analysis to PyVs infecting non-mammalian hosts. We combined biochemical, structural and functional assays to demonstrate that Black Sea Bass (BSB) PyV-LTA is transported into the nucleus by the Importin (IMP)/{beta}1 heterodimer thanks to the recognition of a bipartite cNLS located downstream of the SF3 helicase domain, rather than between the LXCXE motif and the OBD. Such cNLS binds with high affinity to several IMP isoforms by simultaneously interacting with the minor and major binding sites. Substitution of NLS key basic residues abrogating binding to IMP, or co-expression with the well characterized IMP/{beta}1 inhibitor Bimax2 suppressed nuclear localization. Intriguingly a cNLS could be identified in a similar position in LTAs from other PyVs infecting ray-finned fishes, but not cartilaginous fishes, birds or scorpions, where cNLSs were predicted elsewhere. Our study suggests that LTAs from PyVs infecting different non-mammalian hosts might bear cNLS in distinctive positions, possibly reflecting processes of virus-host adaptation.

microbiology↗

bioRxiv2025

Nuclear trafficking of Anelloviridae capsid protein ORF1 reflects modular evolution of subcellular targeting signals

Anelloviridae members are ubiquitous viruses with a small, negative sense, single-stranded DNA genome which is replicated by host cell DNA polymerases. Anelloviruses are postulated to interact with the host cell nuclear transport machinery, however, the lack of reliable cell culture models strongly limits our knowledge regarding Anelloviridae-host interactions. In particular, capsid nuclear import is a largely uncharacterized process. We addressed this by investigating the relationship between host cell nuclear transport receptors (NTRs) and ORF1, the putative capsid protein from torque teno douroucouli virus (TTDoV). We identified the subcellular targeting signals and NTRs responsible for its nucleolar and nuclear localization, and characterized their relative contribution to ORF1 subcellular localization. In the absence of other viral proteins, ORF1 accumulated in the nucleoli. Bioinformatics analysis revealed a putative nuclear localization signal (NLS) within the highly conserved N-terminal arginine rich motif (ARM) ("NLSn", 27-RRWRRRPRRRRRPYR-RRPYRRYGRRRKVRRR-57), and an additional C-terminal NLS ("NLSc", 632-LPPPEKRARWGF-643), which has been specifically acquired by Anelloviridae capsids with larger projection domains. Such NLSs play distinct roles in ORF1 subcellular localization. NLSn features broad importin (IMP) binding affinity yet plays a minor role in nuclear import, being responsible for nucleolar targeting likely through interaction with nucleolar components. NLSc specifically interacts with IMP and is the main driver of active nuclear transport in an IMP/{beta}1-dependent fashion. These findings suggest an evolutionary correlation between the acquisition of progressively larger projection domains and the presence of additional NLSs in Anelloviridae capsids, aimed at maximizing IMP/{beta}1-mediated nuclear import.

microbiology↗