CDK12-mediated transcriptional regulation of noncanonical NF-κB components is essential for signaling.

Members of the family of nuclear factor κB (NF-κB) transcription factors are critical for multiple cellular processes, including regulating innate and adaptive immune responses, cell proliferation, and cell survival. Canonical NF-κB complexes are retained in the cytoplasm by the inhibitory protein IκBα, whereas noncanonical NF-κB complexes are retained by p100. Although activation of canonical NF-κB signaling through the IκBα kinase complex is well studied, few regulators of the NF-κB-inducing kinase (NIK)-dependent processing of noncanonical p100 to p52 and the subsequent nuclear translocation of p52 have been identified.

Investigating small molecules to inhibit germinal center kinase-like kinase (GLK/MAP4K3) upstream of PKCθ phosphorylation: Potential therapy to modulate T cell dependent immunity.

Germinal center kinase-like kinase (GLK, also known as MAP4K3) has been hypothesized to have an effect on key cellular activities, including inflammatory responses. GLK is required for activation of protein kinase C-θ (PKCθ) in T cells. Controlling the activity of T helper cell responses could be valuable for the treatment of autoimmune diseases.

ATP-Competitive MLKL Binders Have No Functional Impact on Necroptosis.

MLKL is a pore forming pseudokinase involved in the final stage of necroptosis, a form of programmed cell death. Its phosphorylation by RIPK3 is necessary for triggering necroptosis but not for triggering apoptosis, which makes it a unique target for pharmacological inhibition to block necroptotic cell death. This mechanism has been described as playing a role in disease progression in neurodegenerative and inflammatory diseases.

Germinal-center kinase-like kinase co-crystal structure reveals a swapped activation loop and C-terminal extension.

Germinal-center kinase-like kinase (GLK, Map4k3), a GCK-I family kinase, plays multiple roles in regulating apoptosis, amino acid sensing, and immune signaling. We describe here the crystal structure of an activation loop mutant of GLK kinase domain bound to an inhibitor. The structure reveals a weakly associated, activation-loop swapped dimer with more than 20 amino acids of ordered density at the carboxy-terminus.

Study of Andes virus entry and neutralization using a pseudovirion system.

Andes virus (ANDV), a member of the Hantavirus genus in the family Bunyaviridae, causes an acute disease characteristic of New-World hantaviruses called hantavirus pulmonary syndrome (HPS). HPS is a highly pathogenic disease with a case-fatality rate of 40%. ANDV is the only hantavirus reported to spread directly from human-to-human.

Impact of methionine oxidation on the binding of human IgG1 to Fc Rn and Fc gamma receptors.

Methionine oxidation commonly occurs in the Fc fragment of therapeutic monoclonal antibodies; however, its impact on antibody function has not been addressed. Using surface plasmon resonance and cell binding assays, we examined the impact of methionine oxidation on the binding of two humanized IgG1 antibodies to Fc gamma receptors (Fc gammaR) and to the neonatal Fc receptor (Fc Rn). A panel of Fc gammaRs, including Fc gammaRI, Fc gammaRIIa-131H, Fc gammaRIIa-131R, Fc gammaRIIb/c, Fc gammaRIII ALF, Fc gammaRIII ALV, and Fc gammaRIIIb was evaluated.

Replication-competent variants of human immunodeficiency virus type 2 lacking the V3 loop exhibit resistance to chemokine receptor antagonists.

Entry of human immunodeficiency virus type 1 (HIV-1) and HIV-2 requires interactions between the envelope glycoprotein (Env) on the virus and CD4 and a chemokine receptor, either CCR5 or CXCR4, on the cell surface. The V3 loop of the HIV gp120 glycoprotein plays a critical role in this process, determining tropism for CCR5- or CXCR4-expressing cells, but details of how V3 interacts with these receptors have not been defined. Using an iterative process of deletion mutagenesis and in vitro adaptation of infectious viruses, variants of HIV-2 were derived that could replicate without V3, either with or without a deletion of the V1/V2 variable loops.

Identification of a novel C-terminal cleavage of Crimean-Congo hemorrhagic fever virus PreGN that leads to generation of an NSM protein.

The structural glycoproteins of Crimean-Congo hemorrhagic fever virus (CCHFV; genus Nairovirus, family Bunyaviridae) are derived through endoproteolytic cleavage of a 1,684-amino-acid M RNA segment-encoded polyprotein. This polyprotein is cotranslationally cleaved into the PreGN and PreGC precursors, which are then cleaved by SKI-1 and a SKI-1-like protease to generate the N termini of GN and GC, respectively. However, the resulting polypeptide defined by the N termini of GN and GC is predicted to be larger (58 kDa) than mature GN (37 kDa).

Development and characterization of a Rift Valley fever virus cell-cell fusion assay using alphavirus replicon vectors.

Rift Valley fever virus (RVFV), a member of the Phlebovirus genus in the Bunyaviridae family, is transmitted by mosquitoes and infects both humans and domestic animals, particularly cattle and sheep. Since primary RVFV strains must be handled in BSL-3+ or BSL-4 facilities, a RVFV cell-cell fusion assay will facilitate the investigation of RVFV glycoprotein function under BSL-2 conditions. As for other members of the Bunyaviridae family, RVFV glycoproteins are targeted to the Golgi, where the virus buds, and are not efficiently delivered to the cell surface.

N-glycans on Nipah virus fusion protein protect against neutralization but reduce membrane fusion and viral entry.

Nipah virus (NiV) is a deadly emerging paramyxovirus. The NiV attachment (NiV-G) and fusion (NiV-F) envelope glycoproteins mediate both syncytium formation and viral entry. Specific N-glycans on paramyxovirus fusion proteins are generally required for proper conformational integrity and biological function.

Two key residues in ephrinB3 are critical for its use as an alternative receptor for Nipah virus.

EphrinB2 was recently discovered as a functional receptor for Nipah virus (NiV), a lethal emerging paramyxovirus. Ephrins constitute a class of homologous ligands for the Eph class of receptor tyrosine kinases and exhibit overlapping expression patterns. Thus, we examined whether other ephrins might serve as alternative receptors for NiV.

Presence of broadly reactive and group-specific neutralizing epitopes on newly described isolates of Crimean-Congo hemorrhagic fever virus.

Crimean-Congo hemorrhagic fever virus (CCHFV), a member of the genus Nairovirus of the family Bunyaviridae, causes severe disease in humans with high rates of mortality. The virus has a tripartite genome composed of a small (S), a medium (M) and a large (L) RNA segment; the M segment encodes the two viral glycoproteins, G(N) and G(C). Whilst relatively few full-length M segment sequences are available, it is apparent that both G(N) and G(C) may exhibit significant sequence diversity.

EphrinB2 is the entry receptor for Nipah virus, an emergent deadly paramyxovirus.

Nipah virus (NiV) is an emergent paramyxovirus that causes fatal encephalitis in up to 70 percent of infected patients, and there is evidence of human-to-human transmission. Endothelial syncytia, comprised of multinucleated giant-endothelial cells, are frequently found in NiV infections, and are mediated by the fusion (F) and attachment (G) envelope glycoproteins. Identification of the receptor for this virus will shed light on the pathobiology of NiV infection, and spur the rational development of effective therapeutics.

Cellular localization and antigenic characterization of crimean-congo hemorrhagic fever virus glycoproteins.

Crimean-Congo hemorrhagic fever virus (CCHFV), a member of the genus Nairovirus of the family Bunyaviridae, causes severe disease with high rates of mortality in humans. The CCHFV M RNA segment encodes the virus glycoproteins G(N) and G(C). To understand the processing and intracellular localization of the CCHFV glycoproteins as well as their neutralization and protection determinants, we produced and characterized monoclonal antibodies (MAbs) specific for both G(N) and G(C).

The 3' end of Norwalk virus mRNA contains determinants that regulate the expression and stability of the viral capsid protein VP1: a novel function for the VP2 protein.

Norwalk virus (NV) is the prototype strain of a group of noncultivable human caliciviruses responsible for epidemic outbreaks of acute gastroenteritis. The capsid protein VP1 is synthesized from a subgenomic RNA that contains two open reading frames (ORFs), ORF2 and ORF3, and the 3' untranslated region (UTR). ORF2 and ORF3 code for the capsid protein (VP1) and a small structural basic protein (VP2), respectively.

Immunogenicity and protective efficacy of rotavirus 2/6-virus-like particles produced by a dual baculovirus expression vector and administered intramuscularly, intranasally, or orally to mice.

Virus-like particles (VLPs) are being evaluated as a candidate rotavirus vaccine. Rotavirus VLPs composed of simian SA11 strain VP2 and VP6 proteins (homologous 2/6-VLPs) were produced by cloning the rotavirus simian SA11 genes 2 and 6 into a single baculovirus transfer vector (pAcAB4). The overall yield of homologous 2/6-VLPs produced with the dual recombinant baculovirus was at least 30-fold higher than that of VLPs composed of bovine RF strain VP2 and simian SA11 strain VP6 (heterologous 2/6-VLPs), produced with single recombinant baculoviruses.

Structural requirements for the assembly of Norwalk virus-like particles.

Norwalk virus (NV) is the prototype strain of a group of human caliciviruses responsible for epidemic outbreaks of acute gastroenteritis. While these viruses do not grow in tissue culture cells or animal models, expression of the capsid protein in insect cells results in the self-assembly of recombinant NV virus-like particles (rNV VLPs) that are morphologically and antigenically similar to native NV. The X-ray structure of the rNV VLPs has revealed that the capsid protein folds into two principal domains: a shell (S) domain and a protruding (P) domain (B.