Qianqian Shao

Tick-borne phenuiviruses include a diverse set of emerging pathogens whose nucleoproteins (NPs) package viral RNA despite extensive sequence divergence across lineages. We examined the NP of Tacheng tick virus 2 (TcTV2), a highly divergent member of the genus Uukuvirus. Phylogenetic analysis of an expanded NP dataset supported the placement of TcTV2 within the broader Uukuvirus lineage, and the AlphaFold model predicted a bilobed core with an electropositive groove resembling those of other phenuivirus NPs. Structural comparisons of the predicted model indicated the greatest variation in the peripheral regions. Recombinant TcTV2 NP co-purified with bacterial RNA and formed heterogeneous ring-like particles, with pentamer-like particles predominating in cryo-EM 2D classes. Truncation of the distal N-terminal region (NP_152–440) did not abolish ring-like particle formation under the tested conditions. Suramin altered the thermal denaturation profile of TcTV2 NP and produced concentration-dependent responses on an NP-loaded BLI sensor in exploratory measurements. Molecular docking and a single 100-ns molecular dynamics simulation suggested a possible suramin-binding mode near the electropositive groove. TcTV2 NP therefore combines a predicted phenuivirus-like core with experimentally observed RNA association and heterogeneous ring-like particle formation, while the computational analyses nominate the electropositive groove as a candidate surface for further ligand interaction studies.

Jumbo phages are a group of tailed bacteriophages with large genomes and capsids. As a prototype of jumbo phage, ΦKZ infects Pseudomonas aeruginosa, a multi-drug-resistant (MDR) opportunistic pathogen leading to acute or chronic infection in immunocompromised individuals. It holds potential to be used as an antimicrobial agent and as a model for uncovering basic phage biology. Although previous low-resolution structural studies have indicated that jumbo phages may have more complicated capsid structures than smaller phages such as HK97, the detailed structures and the assembly mechanism of their capsids remain largely unknown. Here, we report a 3.5-Å-resolution cryo-EM structure of the ΦKZ capsid. The structure unveiled ten minor capsid proteins, with some decorating the outer surface of the capsid and the others forming a complex network attached to the capsid’s inner surface. This network seems to play roles in driving capsid assembly and capsid stabilization. Similar mechanisms of capsid assembly and stabilization are probably employed by many other jumbo viruses.

Epidermal growth factor receptor (EGFR) is expressed in various types of cancer and is associated with the malignant biological behavior of cancer cells. In the present study, the expression of EGFR in hepatocellular carcinoma (HCC) tissues and liver cancer cells was detected by immunohistochemical staining, western blotting and immunofluorescence. Furthermore, a lentivirus was transduced into HepG2 liver cancer cells to knock down EGFR expression. Cell proliferation and migration, and the expression levels of epithelial‑mesenchymal transition (EMT) markers were assessed by EdU staining, Cell Counting Kit‑8, colony formation, wound healing and Transwell assays, and western blotting. The results revealed that EGF/EGFR can mediate EMT through the Akt/glycogen synthase kinase‑3β (GSK-3β)/Snail signaling pathway to promote HepG2 cell proliferation and migration. Inhibition of the activation of the EGFR signaling pathway can help to partially reverse the EMT phenotype, and inhibit the proliferation and migration of HepG2 cells. In conclusion, the EGFR/Akt/GSK-3β/Snail signaling pathway serves an important role in HCC progression, and inhibition of the activation of the EGFR signaling pathway may be a valuable strategy in liver cancer treatment.

Bacteriophage (phage) T4 has served as an extraordinary model to elucidate biological structures and mechanisms. Recent discoveries on the T4 head (capsid) structure, portal vertex, and genome packaging add a significant body of new literature to phage biology. Head structures in unexpanded and expanded conformations show dramatic domain movements, structural remodeling, and a ~70% increase in inner volume while creating high-affinity binding sites for the outer decoration proteins Soc and Hoc. Small changes in intercapsomer interactions modulate angles between capsomer planes, leading to profound alterations in head length. The in situ cryo-EM structure of the symmetry-mismatched portal vertex shows the remarkable structural morphing of local regions of the portal protein, allowing similar interactions with the capsid protein in different structural environments. Conformational changes in these interactions trigger the structural remodeling of capsid protein subunits surrounding the portal vertex, which propagate as a wave of expansion throughout the capsid. A second symmetry mismatch is created when a pentameric packaging motor assembles at the outer “clip” domains of the dodecameric portal vertex. The single-molecule dynamics of the packaging machine suggests a continuous burst mechanism in which the motor subunits adjusted to the shape of the DNA fire ATP hydrolysis, generating speeds as high as 2000 bp/s.

Giant viruses are a large group of viruses that infect many eukaryotes. Although components that do not obey the overall icosahedral symmetry of their capsids have been observed and found to play critical roles in the viral life cycles, identities and high-resolution structures of these components remain unknown. Here, by determining a near-atomic-resolution, five-fold averaged structure of Paramecium bursaria chlorella virus 1, we unexpectedly found the viral capsid possesses up to five major capsid protein variants and a penton protein variant. These variants create varied capsid microenvironments for the associations of fibers, a vesicle, and previously unresolved minor capsid proteins. Our structure reveals the identities and atomic models of the capsid components that do not obey the overall icosahedral symmetry and leads to a model for how these components are assembled and initiate capsid assembly, and this model might be applicable to many other giant viruses.

Many icosahedral viruses assemble proteinaceous precursors called proheads or procapsids. Proheads are metastable structures that undergo a profound structural transition known as expansion that transforms an immature unexpanded head into a mature genome-packaging head. Bacteriophage T4 is a model virus, well studied genetically and biochemically, but its structure determination has been challenging because of its large size and unusually prolate-shaped, ∼1,200-Å-long and ∼860-Å-wide capsid. Here, we report the cryogenic electron microscopy (cryo-EM) structures of T4 capsid in both of its major conformational states: unexpanded at a resolution of 5.1 Å and expanded at a resolution of 3.4 Å. These are among the largest structures deposited in Protein Data Bank to date and provide insights into virus assembly, head length determination, and shell expansion. First, the structures illustrate major domain movements and ∼70% additional gain in inner capsid volume, an essential transformation to contain the entire viral genome. Second, intricate intracapsomer interactions involving a unique insertion domain dramatically change, allowing the capsid subunits to rotate and twist while the capsomers remain fastened at quasi-threefold axes. Third, high-affinity binding sites emerge for a capsid decoration protein that clamps adjacent capsomers, imparting extraordinary structural stability. Fourth, subtle conformational changes at capsomers’ periphery modulate intercapsomer angles between capsomer planes that control capsid length. Finally, conformational changes were observed at the symmetry-mismatched portal vertex, which might be involved in triggering head expansion. These analyses illustrate how small changes in local capsid subunit interactions lead to profound shifts in viral capsid morphology, stability, and volume.

Mosquito viruses cause unpredictable outbreaks of disease. Recently, several unassigned viruses isolated from mosquitoes, including the Omono River virus (OmRV), were identified as totivirus-like viruses, with features similar to those of the Totiviridae family. Most reported members of this family infect fungi or protozoans and lack an extracellular life cycle stage. Here, we identified a new strain of OmRV and determined high-resolution structures for this virus using single-particle cryo-electron microscopy. The structures feature an unexpected protrusion at the five-fold vertex of the capsid. Disassociation of the protrusion could result in several conformational changes in the major capsid. All these structures, together with some biological results, suggest the protrusions’ associations with the extracellular transmission of OmRV. Mosquito is a reservoir of viruses, with a large amount of them perform significant research value. Omono River virus (OmRV) has been isolated from Culex mosquito and is closely related to the family Totiviridae. However, current researches have reported the extracellular transmission ability of OmRV, which is lacked in most members of Totiviridae. In the current study, we isolated a new strain, OmRV-LZ, and obtained its high-resolution cryo-electron microscopy (cryo-EM) structure. Unexpectedly, a protrusion structure has been found located at the five-fold vertex, which is unrecognized in the previous studies. Structural and molecular biological experiments were applied to try to investigate its functions. The results may be helpful to understand the extracellular transmission ability of OmRV-LZ and similar double-stranded RNA (dsRNA) viruses.

Summary: It is well recognized that tissue microenvironments are involved in regulating the development and function of dendritic cells (DC). Oxygen supply, which varies in different tissues, has been accepted as an important microenvironmental factor in regulating the biological functions of several immune cells and as being involved in tumour progression and metastasis. However, little is known about the effect of hypoxia on the biological functions of DC and the effect of these hypoxia‐conditioned DC on tumour metastasis. In this study, we analysed the transcriptional profiles of human monocyte‐derived immature DC (imDC) and mature DC (mDC) cultured under normoxia and hypoxia by microarray, and found a body of potential targets regulating the functions of DC during hypoxia. In addition, the phagocytic ability of hypoxic imDC markedly decreased compared with that of normoxic imDC. Importantly, hypoxic DC poorly induced the proliferation of allogeneic T cells, but polarized allogeneic CD4⁺ naive T cells into a T helper type 2 (Th2) response. Moreover, hypoxic DC secreted large amounts of osteopontin, which were responsible for the enhanced migration of tumour cells. Therefore, our study provides new insights into the biological functions of DC under hypoxic conditions and one of mechanisms underlying tumour immune escape during hypoxia.