Surface microlayer-mediated virome dissemination in the Central Arctic.

Background: Aquatic viruses act as key players in shaping microbial communities. In polar environments, they face significant challenges such as limited host availability and harsh conditions. However, due to the restricted accessibility of these ecosystems, our understanding of viral diversity, abundance, adaptations, and host interactions remains limited.

Structure-based discovery of first inhibitors targeting the helicase activity of human PIF1.

PIF1 is a conserved helicase and G4 DNA binding and unwinding enzyme, with roles in genome stability. Human PIF1 (hPIF1) is poorly understood, but its functions can become critical for tumour cell survival during oncogene-driven replication stress. Here we report the discovery, via an X-ray crystallographic fragment screen (XChem), of hPIF1 DNA binding and unwinding inhibitors.

Structural basis for DNA recognition by a viral genome-packaging machine.

DNA recognition is critical for assembly of double-stranded DNA viruses, particularly for the initiation of packaging the viral genome into the capsid. The key component that recognizes viral DNA is the small terminase protein. Despite prior studies, the molecular mechanism for DNA recognition remained elusive.

Viral Genomic DNA Packaging Machinery.

Tailed double-stranded DNA bacteriophage employs a protein terminase motor to package their genome into a preformed protein shell-a system shared with eukaryotic dsDNA viruses such as herpesviruses. DNA packaging motor proteins represent excellent targets for antiviral therapy, with Letermovir, which binds Cytomegalovirus terminase, already licensed as an effective prophylaxis. In the realm of bacterial viruses, these DNA packaging motors comprise three protein constituents: the portal protein, small terminase and large terminase.

A stargate mechanism of genome delivery unveiled by cryogenic electron tomography.

Single-stranded DNA bacteriophages of the family are major components of the global virosphere. Microviruses are highly abundant in aquatic ecosystems and are prominent members of the mammalian gut microbiome, where their diversity has been linked to various chronic health disorders. Despite the clear importance of microviruses, little is known about the molecular mechanism of host infection.

Jorvik: A membrane-containing phage that will likely found a new family within Vinavirales.

Although membrane-containing dsDNA bacterial viruses are some of the most prevalent predators in aquatic environments, we know little about how they function due to their intractability in the laboratory. Here, we have identified and thoroughly characterized a new type of membrane-containing bacteriophage, Jorvik, that infects the freshwater mixotrophic model bacterium . Jorvik is extremely virulent, can persist in the host integrated into the RuBisCo operon and encodes two experimentally verified cell wall hydrolases.

Ecogenomics and cultivation reveal distinctive viral-bacterial communities in the surface microlayer of a Baltic Sea slick.

Visible surface films, termed slicks, can extensively cover freshwater and marine ecosystems, with coastal regions being particularly susceptible to their presence. The sea-surface microlayer (SML), the upper 1-mm at the air-water interface in slicks (herein slick SML) harbors a distinctive bacterial community, but generally little is known about SML viruses. Using flow cytometry, metagenomics, and cultivation, we characterized viruses and bacteria in a brackish slick SML in comparison to non-slick SML as well as seawater below slick and non-slick areas (subsurface water = SSW).

Structure of HK97 small terminase:DNA complex unveils a novel DNA binding mechanism by a circular protein.

DNA recognition is critical for assembly of double-stranded DNA viruses, in particular for the initiation of packaging the viral genome into the capsid. DNA packaging has been extensively studied for three archetypal bacteriophage systems: , and phi29. We identified the minimal site within the region of bacteriophage HK97 specifically recognised by the small terminase and determined a cryoEM structure for the small terminase:DNA complex.

Insights into a viral motor: the structure of the HK97 packaging termination assembly.

Double-stranded DNA viruses utilise machinery, made of terminase proteins, to package viral DNA into the capsid. For cos bacteriophage, a defined signal, recognised by small terminase, flanks each genome unit. Here we present the first structural data for a cos virus DNA packaging motor, assembled from the bacteriophage HK97 terminase proteins, procapsids encompassing the portal protein, and DNA containing a cos site.

Structural atlas of a human gut crassvirus.

CrAssphage and related viruses of the order Crassvirales (hereafter referred to as crassviruses) were originally discovered by cross-assembly of metagenomic sequences. They are the most abundant viruses in the human gut, are found in the majority of individual gut viromes, and account for up to 95% of the viral sequences in some individuals. Crassviruses are likely to have major roles in shaping the composition and functionality of the human microbiome, but the structures and roles of most of the virally encoded proteins are unknown, with only generic predictions resulting from bioinformatic analyses.

High-Voltage Biomolecular Sensing Using a Bacteriophage Portal Protein Covalently Immobilized within a Solid-State Nanopore.

The application of nanopores as label-free, single-molecule biosensors for electrical or optical probing of structural features in biomolecules has been widely explored. While biological nanopores (membrane proteins and bacteriophage portal proteins) and solid-state nanopores (thin films and two-dimensional materials) have been extensively employed, the third class of nanopores known as hybrid nanopores, where an artificial membrane substitutes the organic support membrane of proteins, has been only sparsely studied due to challenges in implementation. portal protein contains a natural DNA pore that is used by viruses for filling their capsid with viral genomic DNA.

Human 14-3-3 Proteins Site-selectively Bind the Mutational Hotspot Region of SARS-CoV-2 Nucleoprotein Modulating its Phosphoregulation.

Phosphorylation of SARS-CoV-2 nucleoprotein recruits human cytosolic 14-3-3 proteins playing a well-recognized role in replication of many viruses. Here we use genetic code expansion to demonstrate that 14-3-3 binding is triggered by phosphorylation of SARS-CoV-2 nucleoprotein at either of two pseudo-repeats centered at Ser197 and Thr205. According to fluorescence anisotropy measurements, the pT205-motif,presentin SARS-CoV-2 but not in SARS-CoV, is preferred over the pS197-motif by all seven human 14-3-3 isoforms, which collectively display an unforeseen pT205/pS197 peptide binding selectivity hierarchy.

Structural basis of DNA packaging by a ring-type ATPase from an archetypal viral system.

Many essential cellular processes rely on substrate rotation or translocation by a multi-subunit, ring-type NTPase. A large number of double-stranded DNA viruses, including tailed bacteriophages and herpes viruses, use a homomeric ring ATPase to processively translocate viral genomic DNA into procapsids during assembly. Our current understanding of viral DNA packaging comes from three archetypal bacteriophage systems: cos, pac and phi29.

Prevalence of congenital cryptorchidism in Estonia.

Background: Cryptorchidism is one of the most common urogenital malformations. Cryptorchidism prevalence varies greatly in different countries and populations. The aim of the current study was to determine and analyse cryptorchidism prevalence in Estonia.

CryoEM structure of the Nipah virus nucleocapsid assembly.

Nipah and its close relative Hendra are highly pathogenic zoonotic viruses, storing their ssRNA genome in a helical nucleocapsid assembly formed by the N protein, a major viral immunogen. Here, we report the first cryoEM structure for a Henipavirus RNA-bound nucleocapsid assembly, at 3.5 Å resolution.

The Mechanism of SARS-CoV-2 Nucleocapsid Protein Recognition by the Human 14-3-3 Proteins.

The coronavirus nucleocapsid protein (N) controls viral genome packaging and contains numerous phosphorylation sites located within unstructured regions. Binding of phosphorylated SARS-CoV N to the host 14-3-3 protein in the cytoplasm was reported to regulate nucleocytoplasmic N shuttling. All seven isoforms of the human 14-3-3 are abundantly present in tissues vulnerable to SARS-CoV-2, where N can constitute up to ~1% of expressed proteins during infection.

Broad and strong memory CD4 and CD8 T cells induced by SARS-CoV-2 in UK convalescent individuals following COVID-19.

The development of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) vaccines and therapeutics will depend on understanding viral immunity. We studied T cell memory in 42 patients following recovery from COVID-19 (28 with mild disease and 14 with severe disease) and 16 unexposed donors, using interferon-γ-based assays with peptides spanning SARS-CoV-2 except ORF1. The breadth and magnitude of T cell responses were significantly higher in severe as compared with mild cases.

Molecular basis for the recognition of steroidogenic acute regulatory protein by the 14-3-3 protein family.

Steroidogenesis in adrenals and gonads starts from cholesterol transport to mitochondria. This is mediated by the steroidogenic acute regulatory protein (STARD1), containing a mitochondrial import sequence followed by a cholesterol-binding START domain. Although mutations in this protein have been linked to lipoid congenital adrenal hyperplasia (LCAH), the mechanism of steroidogenesis regulation by STARD1 remains debatable.

Cryo-EM structure in situ reveals a molecular switch that safeguards virus against genome loss.

The portal protein is a key component of many double-stranded DNA viruses, governing capsid assembly and genome packaging. Twelve subunits of the portal protein define a tunnel, through which DNA is translocated into the capsid. It is unknown how the portal protein functions as a gatekeeper, preventing DNA slippage, whilst allowing its passage into the capsid, and how these processes are controlled.

Diversity and Host Interactions Among Virulent and Temperate Baltic Sea Phages.

Viruses in aquatic environments play a key role in microbial population dynamics and nutrient cycling. In particular, bacteria of the phylum are known to participate in recycling algal blooms. Studies of phage-host interactions involving this phylum are hence important to understand the processes shaping bacterial and viral communities in the ocean as well as nutrient cycling.

Cryo-EM structure and in vitro DNA packaging of a thermophilic virus with supersized T=7 capsids.

Double-stranded DNA viruses, including bacteriophages and herpesviruses, package their genomes into preformed capsids, using ATP-driven motors. Seeking to advance structural and mechanistic understanding, we established in vitro packaging for a thermostable bacteriophage, P23-45 of Both the unexpanded procapsid and the expanded mature capsid can package DNA in the presence of packaging ATPase over the 20 °C to 70 °C temperature range, with optimum activity at 50 °C to 65 °C. Cryo-EM reconstructions for the mature and immature capsids at 3.

Structural and functional analysis of the nucleotide and DNA binding activities of the human PIF1 helicase.

Pif1 is a multifunctional helicase and DNA processing enzyme that has roles in genome stability. The enzyme is conserved in eukaryotes and also found in some prokaryotes. The functions of human PIF1 (hPIF1) are also critical for survival of certain tumour cell lines during replication stress, making it an important target for cancer therapy.

Thermostable virus portal proteins as reprogrammable adapters for solid-state nanopore sensors.

Nanopore-based sensors are advancing the sensitivity and selectivity of single-molecule detection in molecular medicine and biotechnology. Current electrical sensing devices are based on either membrane protein pores supported in planar lipid bilayers or solid-state (SS) pores fabricated in thin metallic membranes. While both types of nanosensors have been used in a variety of applications, each has inherent disadvantages that limit its use.

Porphyrin-Assisted Docking of a Thermophage Portal Protein into Lipid Bilayers: Nanopore Engineering and Characterization.

Nanopore-based sensors for nucleic acid sequencing and single-molecule detection typically employ pore-forming membrane proteins with hydrophobic external surfaces, suitable for insertion into a lipid bilayer. In contrast, hydrophilic pore-containing molecules, such as DNA origami, have been shown to require chemical modification to favor insertion into a lipid environment. In this work, we describe a strategy for inserting polar proteins with an inner pore into lipid membranes, focusing here on a circular 12-subunit assembly of the thermophage G20c portal protein.