Dehydrocoupling of Phosphine-Borane Adducts Under Ambient Conditions Using Aminoboranes as Hydrogen Acceptors.

We report on the reactivity of aminoboranes (RN=BH; R = iPr, Et, Me) with phosphine-borane adducts (PhR'PH ⋅ BH; R' = H, Ph): Sufficiently sterically unencumbered aminoboranes can accept hydrogen from phosphine-borane adducts. The hydrogen transfer results in the formation of amine-borane adducts (RNH ⋅ BH) and transient phosphinoboranes (PhR'P-BH) in situ. These phosphinoboranes undergo subsequent reactivity to yield either polyphosphinoborane, [PhPH-BH], or the linear dimer, PhPH ⋅ BH-PhP ⋅ BH.

Polyphosphinoborane Block Copolymer Synthesis Using Catalytic Reversible Chain-Transfer Dehydropolymerization.

An amphiphilic block copolymer of polyphosphinoborane has been prepared by a mechanism-led strategy of the sequential catalytic dehydropolymerization of precursor monomers, H B ⋅ PRH (R=Ph, n-hexyl), using the simple pre-catalyst [Rh(Ph PCH CH PPh ) ]Cl. Speciation, mechanism and polymer chain growth studies support a step-growth process where reversible chain transfer occurs, i.e.

Transition-Metal-Free Dehydropolymerization of Phosphine-Boranes at Ambient Temperature.

Stoichiometric reaction of phosphine-borane adducts RR'PH⋅BH (R=Ph, R'=H, Ph, Et, and R=R'= Bu) with the strong acid HNTf (Tf=SO CF ) leads to H elimination and the formation of the triflimido derivatives, RR'PH⋅BH (NTf ). Subsequent deprotonation by using bases, such as diisopropylethylamine or the carbene IPr (IPr=N,N'-bis(2,6-diisopropylphenyl)imidazol-2-ylidene), led to the formation of P-mono- or -disubstituted polyphosphinoboranes [RR'P-BH ] . Evidence for the intermediacy of transient phosphinoborane monomers, RR'PBH , was provided by trapping reactions.

Dysregulation of Cellular VRK1, BAF, and Innate Immune Signaling by the Vaccinia Virus B12 Pseudokinase.

Poxvirus proteins remodel signaling throughout the cell by targeting host enzymes for inhibition and redirection. Recently, it was discovered that early in infection the vaccinia virus (VACV) B12 pseudokinase copurifies with the cellular kinase VRK1, a proviral factor, in the nucleus. Although the formation of this complex correlates with inhibition of cytoplasmic VACV DNA replication and likely has other downstream signaling consequences, the molecular mechanisms involved are poorly understood.

Calix[4]pyrrolato Aluminate Catalyzes the Dehydrocoupling of Phenylphosphine Borane to High Molar Weight Polymers.

High molar weight polyphosphinoboranes represent materials with auspicious properties, but their preparation requires transition metal-based catalysts. Here, calix[4]pyrrolato aluminate is shown to induce the dehydropolymerization of phosphine boranes to high molar mass polyphosphinoboranes (up to M =43 000 Da). Combined GPC and P DOSY NMR spectroscopic analyses, quantum chemical computations, and stoichiometric reactions disclose a P-H bond activation by the cooperative action of the square-planar aluminate and the electron-rich ligand framework.

Vaccinia Virus Arrests and Shifts the Cell Cycle.

Modulation of the host cell cycle is a common strategy used by viruses to create a pro-replicative environment. To facilitate viral genome replication, vaccinia virus (VACV) has been reported to alter cell cycle regulation and trigger the host cell DNA damage response. However, the cellular factors and viral effectors that mediate these changes remain unknown.

The Vaccinia Virus B12 Pseudokinase Represses Viral Replication via Interaction with the Cellular Kinase VRK1 and Activation of the Antiviral Effector BAF.

The poxviral B1 and B12 proteins are a homologous kinase-pseudokinase pair, which modulates a shared host pathway governing viral DNA replication and antiviral defense. While the molecular mechanisms involved are incompletely understood, B1 and B12 seem to intersect with signaling processes mediated by their cellular homologs termed the vaccinia-related kinases (VRKs). In this study, we expand upon our previous characterization of the B1-B12 signaling axis to gain insights into B12 function.

The Vaccinia Virus (VACV) B1 and Cellular VRK2 Kinases Promote VACV Replication Factory Formation through Phosphorylation-Dependent Inhibition of VACV B12.

Comparative examination of viral and host protein homologs reveals novel mechanisms governing downstream signaling effectors of both cellular and viral origin. The vaccinia virus B1 protein kinase is involved in promoting multiple facets of the virus life cycle and is a homolog of three conserved cellular enzymes called vaccinia virus-related kinases (VRKs). Recent evidence indicates that B1 and VRK2 mediate a common pathway that is largely uncharacterized but appears independent of previous VRK substrates.

Generation of Vaccinia Virus Gene Deletion Mutants Using Complementing Cell Lines.

This protocol describes how to couple two techniques, the generation of complementing cells lines and production of viral deletion mutants, to rapidly construct novel tools for poxvirus analysis. Specifically, the production and utilization of a complementing cell line expressing a poxvirus gene of interest are critical for the generation of poxvirus mutants in which essential genes are disrupted. Complementing cells are also valuable for the characterization of vaccinia genes in the absence of infection.

A poxvirus pseudokinase represses viral DNA replication via a pathway antagonized by its paralog kinase.

Poxviruses employ sophisticated, but incompletely understood, signaling pathways that engage cellular defense mechanisms and simultaneously ensure viral factors are modulated properly. For example, the vaccinia B1 protein kinase plays a vital role in inactivating the cellular antiviral factor BAF, and likely orchestrates other pathways as well. In this study, we utilized experimental evolution of a B1 deletion virus to perform an unbiased search for suppressor mutations and identify novel pathways involving B1.

A cell death assay for assessing the mitochondrial targeting of proteins.

The mitochondrial proteome comprises 1000 to 1500 proteins, in addition to proteins for which the mitochondrial localization is uncertain. About 800 diseases have been linked with mutations in mitochondrial proteins. We devised a cell survival assay for assessing the mitochondrial localization in a high-throughput format.

Diverse silver(i) coordination chemistry with cyclic selenourea ligands.

The coordination chemistry of two selenourea ligands (SeIMes and SeIPr) towards silver(i) triflate and silver(i) nitrate was investigated. Two aggregation modes were observed in the solid state, strongly influenced by the size of the aromatic substituents on the ligand. With mesityl groups, selenium-bridged bimetallic motifs [AgX(SeIMes)] were obtained, while for the bulkier diisopropylphenyl groups ion-separated species of formulae [Ag(SeIPr)][X] were obtained.

Deletion of the Vaccinia Virus B1 Kinase Reveals Essential Functions of This Enzyme Complemented Partly by the Homologous Cellular Kinase VRK2.

The vaccinia virus B1 kinase is highly conserved among poxviruses and is essential for the viral life cycle. B1 exhibits a remarkable degree of similarity to vaccinia virus-related kinases (VRKs), a family of cellular kinases, suggesting that the viral enzyme has evolved to mimic VRK activity. Indeed, B1 and VRKs have been demonstrated to target a shared substrate, the DNA binding protein BAF, elucidating a signaling pathway important for both mitosis and the antiviral response.

The Barrier to Autointegration Factor: Interlocking Antiviral Defense with Genome Maintenance.

Intrinsic defenses targeting foreign DNA are one facet of the cellular armament tasked with protecting host genomic integrity. The DNA binding protein BAF (barrier to autointegration factor) contributes to multiple aspects of genome maintenance and intercepts retrovirus, poxvirus, and herpesvirus genomes during infection. In this gem, we discuss the unique position BAF occupies at the virus-host interface and how both viral and cellular mechanisms may regulate its capacity to act as a pro- or antiviral effector targeting viral DNA.

Vaccinia Virus B1 Kinase Is Required for Postreplicative Stages of the Viral Life Cycle in a BAF-Independent Manner in U2OS Cells.

Unlabelled: The vaccinia virus B1R gene encodes a highly conserved protein kinase that is essential for the poxviral life cycle. As demonstrated in many cell types, B1 plays a critical role during viral DNA replication when it inactivates the cellular host defense effector barrier to autointegration factor (BAF or BANF1). To better understand the role of B1 during infection, we have characterized the growth of a B1-deficient temperature-sensitive mutant virus (Cts2 virus) in U2OS osteosarcoma cells.

Barrier to Autointegration Factor (BANF1): interwoven roles in nuclear structure, genome integrity, innate immunity, stress responses and progeria.

The Barrier to Autointegration Factor (BAF or BANF1) is an abundant, highly conserved DNA binding protein. BAF is involved in multiple pathways including mitosis, nuclear assembly, viral infection, chromatin and gene regulation and the DNA damage response. BAF is also essential for early development in metazoans and relevant to human physiology; BANF1 mutations cause a progeroid syndrome, placing BAF within the laminopathy disease spectrum.

Barrier to auto integration factor becomes dephosphorylated during HSV-1 Infection and Can Act as a host defense by impairing viral DNA replication and gene expression.

BAF (Barrier to Autointegration Factor) is a highly conserved DNA binding protein that senses poxviral DNA in the cytoplasm and tightly binds to the viral genome to interfere with DNA replication and transcription. To counteract BAF, a poxviral-encoded protein kinase phosphorylates BAF, which renders BAF unable to bind DNA and allows efficient viral replication to occur. Herein, we examined how BAF phosphorylation is affected by herpes simplex virus type 1 (HSV-1) infection and tested the ability of BAF to interfere with HSV-1 productive infection.

Bovine herpesvirus 1 productive infection stimulates inflammasome formation and caspase 1 activity.

Bovine herpesvirus 1 (BoHV-1), a significant viral pathogen of cattle, causes inflammation in affected tissue during acute infection. Consequently, we tested whether productively infected bovine cells stimulate inflammasome formation. Expression of two components required for inflammasome formation, the DNA sensor IFI16 (gamma-interferon-inducible protein 16) and NLRP3 (NOD-like receptor family, pyrin domain containing 3), were induced in bovine kidney cells by eight hours after infection.

Cell- and virus-mediated regulation of the barrier-to-autointegration factor's phosphorylation state controls its DNA binding, dimerization, subcellular localization, and antipoxviral activity.

Unlabelled: Barrier-to-autointegration factor (BAF) is a DNA binding protein with multiple cellular functions, including the ability to act as a potent defense against vaccinia virus infection. This antiviral function involves BAF's ability to condense double-stranded DNA and subsequently prevent viral DNA replication. In recent years, it has become increasingly evident that dynamic phosphorylation involving the vaccinia virus B1 kinase and cellular enzymes is likely a key regulator of multiple BAF functions; however, the precise mechanisms are poorly understood.

Barrier to autointegration factor (BAF) inhibits vaccinia virus intermediate transcription in the absence of the viral B1 kinase.

Barrier to autointegration factor (BAF/BANF1) is a cellular DNA-binding protein found in the nucleus and cytoplasm. Cytoplasmic BAF binds to foreign DNA and can act as a defense against vaccinia DNA replication. To evade BAF, vaccinia expresses the B1 kinase, which phosphorylates BAF and blocks its ability to bind DNA.

Molecular genetic and biochemical characterization of the vaccinia virus I3 protein, the replicative single-stranded DNA binding protein.

Vaccinia virus, the prototypic poxvirus, efficiently and faithfully replicates its ∼200-kb DNA genome within the cytoplasm of infected cells. This intracellular localization dictates that vaccinia virus encodes most, if not all, of its own DNA replication machinery. Included in the repertoire of viral replication proteins is the I3 protein, which binds to single-stranded DNA (ssDNA) with great specificity and stability and has been presumed to be the replicative ssDNA binding protein (SSB).

Molecular characterization of the host defense activity of the barrier to autointegration factor against vaccinia virus.

The barrier to autointegration factor (BAF) is an essential cellular protein with functions in mitotic nuclear reassembly, retroviral preintegration complex stability, and transcriptional regulation. Molecular properties of BAF include the ability to bind double-stranded DNA in a sequence-independent manner, homodimerize, and bind proteins containing a LEM domain. These capabilities allow BAF to compact DNA and assemble higher-order nucleoprotein complexes, the nature of which is poorly understood.

Banf1 is required to maintain the self-renewal of both mouse and human embryonic stem cells.

Self-renewal is a complex biological process necessary for maintaining the pluripotency of embryonic stem cells (ESCs). Recent studies have used global proteomic techniques to identify proteins that associate with the master regulators Oct4, Nanog and Sox2 in ESCs or in ESCs during the early stages of differentiation. Through an unbiased proteomic screen, Banf1 was identified as a Sox2-associated protein.

Mice deficient in the serine/threonine protein kinase VRK1 are infertile due to a progressive loss of spermatogonia.

The VRK1 protein kinase has been implicated as a pro-proliferative factor. Genetic analyses of mutant alleles of the Drosophila and Caenorhabditis elegans VRK1 homologs have revealed phenotypes ranging from embryonic lethality to mitotic and meiotic defects with resultant sterility. Herein, we describe the first genetic analysis of murine VRK1.

Poxviral B1 kinase overcomes barrier to autointegration factor, a host defense against virus replication.

Barrier to autointegration factor (BAF) is a DNA-binding protein found in the nucleus and cytoplasm of eukaryotic cells that functions to establish nuclear architecture during mitosis. Herein, we demonstrate a cytoplasmic role for BAF in host defense during poxviral infections. Vaccinia is the prototypic poxvirus, a family of DNA viruses that replicate exclusively in the cytoplasm of infected cells.