Loss of reduces adipogenesis and improves insulin sensitivity in mouse and human adipocytes.

There are multiple independent genetic signals at the () locus associated with type 2 diabetes risk, fasting glucose, ectopic fat, height, and bone mineral density. We have previously shown that loss of in pancreatic beta cells reduces insulin content and impairs islet cell development and function. However, RREB1 is a widely expressed transcription factor and the metabolic impact of RREB1 loss remains unknown.

PAX4 loss of function increases diabetes risk by altering human pancreatic endocrine cell development.

The coding variant (p.Arg192His) in the transcription factor PAX4 is associated with an altered risk for type 2 diabetes (T2D) in East Asian populations. In mice, Pax4 is essential for beta cell formation but its role on human beta cell development and/or function is unknown.

Loss of RREB1 in pancreatic beta cells reduces cellular insulin content and affects endocrine cell gene expression.

Aims/hypothesis: Genome-wide studies have uncovered multiple independent signals at the RREB1 locus associated with altered type 2 diabetes risk and related glycaemic traits. However, little is known about the function of the zinc finger transcription factor Ras-responsive element binding protein 1 (RREB1) in glucose homeostasis or how changes in its expression and/or function influence diabetes risk.

Methods: A zebrafish model lacking rreb1a and rreb1b was used to study the effect of RREB1 loss in vivo.

Multi-ancestry genetic study of type 2 diabetes highlights the power of diverse populations for discovery and translation.

We assembled an ancestrally diverse collection of genome-wide association studies (GWAS) of type 2 diabetes (T2D) in 180,834 affected individuals and 1,159,055 controls (48.9% non-European descent) through the Diabetes Meta-Analysis of Trans-Ethnic association studies (DIAMANTE) Consortium. Multi-ancestry GWAS meta-analysis identified 237 loci attaining stringent genome-wide significance (P < 5 × 10), which were delineated to 338 distinct association signals.

A CRISPR/Cas9 genome editing pipeline in the EndoC-βH1 cell line to study genes implicated in beta cell function.

Type 2 diabetes (T2D) is a global pandemic with a strong genetic component, but most causal genes influencing the disease risk remain unknown. It is clear, however, that the pancreatic beta cell is central to T2D pathogenesis. gene-knockout (KO) models to study T2D risk genes have so far focused on rodent beta cells.

Deep learning models predict regulatory variants in pancreatic islets and refine type 2 diabetes association signals.

Genome-wide association analyses have uncovered multiple genomic regions associated with T2D, but identification of the causal variants at these remains a challenge. There is growing interest in the potential of deep learning models - which predict epigenome features from DNA sequence - to support inference concerning the regulatory effects of disease-associated variants. Here, we evaluate the advantages of training convolutional neural network (CNN) models on a broad set of epigenomic features collected in a single disease-relevant tissue - pancreatic islets in the case of type 2 diabetes (T2D) - as opposed to models trained on multiple human tissues.

Loss of ZnT8 function protects against diabetes by enhanced insulin secretion.

A rare loss-of-function allele p.Arg138* in SLC30A8 encoding the zinc transporter 8 (ZnT8), which is enriched in Western Finland, protects against type 2 diabetes (T2D). We recruited relatives of the identified carriers and showed that protection was associated with better insulin secretion due to enhanced glucose responsiveness and proinsulin conversion, particularly when compared with individuals matched for the genotype of a common T2D-risk allele in SLC30A8, p.

Efficacy of arginine depletion by ADI-PEG20 in an intracranial model of GBM.

Glioblastoma multiforme (GBM) remains a cancer with a poor prognosis and few effective therapeutic options. Successful medical management of GBM is limited by the restricted access of drugs to the central nervous system (CNS) caused by the blood brain barrier (BBB). We previously showed that a subset of GBM are arginine auxotrophic because of transcriptional silencing of ASS1 and/or ASL and are sensitive to pegylated arginine deiminase (ADI-PEG20).

Systems Analysis of Protein Fatty Acylation in Herpes Simplex Virus-Infected Cells Using Chemical Proteomics.

Protein fatty acylation regulates diverse aspects of cellular function and organization and plays a key role in host immune responses to infection. Acylation also modulates the function and localization of virus-encoded proteins. Here, we employ chemical proteomics tools, bio-orthogonal probes, and capture reagents to study myristoylation and palmitoylation during infection with herpes simplex virus (HSV).

Phosphorylation and SCF-mediated degradation regulate CREB-H transcription of metabolic targets.

CREB‑H, an endoplasmic reticulum-anchored transcription factor, plays a key role in regulating secretion and in metabolic and inflammatory pathways, but how its activity is modulated remains unclear. We examined processing of the nuclear active form and identified a motif around S87-S90 with homology to DSG-type phosphodegrons. We show that this region is subject to multiple phosphorylations, which regulate CREB-H stability by targeting it to the SCF(Fbw1a) E3 ubiquitin ligase.

Polarized cell migration during cell-to-cell transmission of herpes simplex virus in human skin keratinocytes.

In addition to transmission involving extracellular free particles, a generally accepted model of virus propagation is one wherein virus replicates in one cell, producing infectious particles that transmit to the next cell via cell junctions or induced polarized contacts. This mechanism of spread is especially important in the presence of neutralizing antibody, and the concept underpins analysis of virus spread, plaque size, viral and host functions, and general mechanisms of virus propagation. Here, we demonstrate a novel process involved in cell-to-cell transmission of herpes simplex virus (HSV) in human skin cells that has not previously been appreciated.

Host-range restriction of vaccinia virus E3L deletion mutant can be overcome in vitro, but not in vivo, by expression of the influenza virus NS1 protein.

During the last decades, research focused on vaccinia virus (VACV) pathogenesis has been intensified prompted by its potential beneficial application as a vector for vaccine development and anti-cancer therapies, but also due to the fear of its potential use as a bio-terrorism threat. Recombinant viruses lacking a type I interferon (IFN) antagonist are attenuated and hence good vaccine candidates. However, vaccine virus growth requires production in IFN-deficient systems, and thus viral IFN antagonists that are active in vitro, yet not in vivo, are of great value.

Differing roles of inner tegument proteins pUL36 and pUL37 during entry of herpes simplex virus type 1.

Studies with herpes simplex virus type 1 (HSV-1) have shown that secondary envelopment and virus release are blocked in mutants deleted for the tegument protein gene UL36 or UL37, leading to the accumulation of DNA-containing capsids in the cytoplasm of infected cells. The failure to assemble infectious virions has meant that the roles of these genes in the initial stages of infection could not be investigated. To circumvent this, cells infected at a low multiplicity were fused to form syncytia, thereby allowing capsids released from infected nuclei access to uninfected nuclei without having to cross a plasma membrane.

Activation mechanism of a noncanonical RNA-dependent RNA polymerase.

Two lineages of viral RNA-dependent RNA polymerases (RDRPs) differing in the organization (canonical vs. noncanonical) of the palm subdomain have been identified. Phylogenetic analyses indicate that both lineages diverged at a very early stage of the evolution of the enzyme [Gorbalenya AE, Pringle FM, Zeddam JL, Luke BT, Cameron CE, Kalmakoff J, Hanzlik TN, Gordon KH, Ward VK (2002) J Mol Biol 324:47-62].

The 2.6-Angstrom structure of infectious bursal disease virus-derived T=1 particles reveals new stabilizing elements of the virus capsid.

Infectious bursal disease virus (IBDV), a member of the Birnaviridae family, is a double-stranded RNA virus that causes a highly contagious disease in young chickens leading to significant economic losses in the poultry industry. The VP2 protein, the only structural component of the IBDV icosahedral capsid, spontaneously assembles into T=1 subviral particles (SVP) when individually expressed as a chimeric gene. We have determined the crystal structure of the T=1 SVP to 2.

Intracellular interference of infectious bursal disease virus.

A search for dominant-negative mutant polypeptides hampering infectious bursal disease virus (IBDV) replication has been undertaken. We have found that expression of a mutant version of the VP3 structural polypeptide known as VP3/M3, partially lacking the domain responsible for the interaction with the virus-encoded RNA polymerase, efficiently interferes with the IBDV replication cycle. Transformed cells stably expressing VP3/M3 show a significant reduction (up to 96%) in their ability to support IBDV growth.

Improving recombinant MVA immune responses: potentiation of the immune responses to HIV-1 with MVA and DNA vectors expressing Env and the cytokines IL-12 and IFN-gamma.

Recombinants based on vaccinia virus vectors, especially on the highly attenuated modified vaccinia virus Ankara (MVA) strain, are now being tested in clinical trials for safety and immunogenicity, using prime/boost heterologous regimes of vaccination. Due to the limited replication capacity of MVA, it is necessary to develop procedures that can enhance the specific cellular immune responses to the recombinant antigen delivered by the MVA vector. In this investigation, we have characterized the systemic immune responses in BALB/c mice using interferon-gamma (IFN-gamma) or interleukin-12 (IL-12) in an adjuvant-like manner elicited by MVA recombinants or naked DNA vectors expressing one of those cytokines in combination with the human immunodeficiency virus type 1 (HIV-1) envelope (Env) as antigen.

Efficient CD8+ T cell response to the HIV-env V3 loop epitope from multiple virus isolates by a DNA prime/vaccinia virus boost (rWR and rMVA strains) immunization regime and enhancement by the cytokine IFN-gamma.

The cytotoxic T-lymphocyte response (CTL) has been shown to be determinant in the clearance of many viral infections and hence, vaccine candidates against AIDS are designed to enhance this arm of the immune system. In this study, we have analyzed the antigen specific immune responses triggered in mice by different combinations of vaccine vehicles expressing the multiepitope polypeptide TAB13. This chimeric protein contains the V3 region of the gp120 from eight different HIV-1 isolates and was efficiently expressed by a DNA vector (DNA-TAB), and also by vaccinia virus recombinants (rVV) based either on the attenuated modified vaccinia virus Ankara (MVA-TAB) or Western Reserve (VV-TAB) strains.

The C-terminal domain of the pVP2 precursor is essential for the interaction between VP2 and VP3, the capsid polypeptides of infectious bursal disease virus.

The interaction between the infectious bursal disease virus (IBDV) capsid proteins VP2 and VP3 has been analyzed in vivo using baculovirus expression vectors. Data presented here demonstrate that the 71-amino acid C-terminal-specific domain of pVP2, the VP2 precursor, is essential for the establishment of the VP2-VP3 interaction. Additionally, we show that coexpression of the pVP2 and VP3 polypeptides from independent genes results in the assembly of virus-like particles (VLPs).

The oligomerization domain of VP3, the scaffolding protein of infectious bursal disease virus, plays a critical role in capsid assembly.

Infectious bursal disease virus (IBDV) capsids are formed by a single protein layer containing three polypeptides, pVP2, VP2, and VP3. Here, we show that the VP3 protein synthesized in insect cells, either after expression of the complete polyprotein or from a VP3 gene construct, is proteolytically degraded, leading to the accumulation of product lacking the 13 C-terminal residues. This finding led to identification of the VP3 oligomerization domain within a 24-amino-acid stretch near the C-terminal end of the polypeptide, partially overlapping the VP1 binding domain.

In vitro survival and germination of Candida albicans in the presence of nitrogen compounds.

The in vitro effect of nitric oxide (NO) and nitrite on blastoconidia and hyphae of Candida albicans was studied. Sodium nitroprusside (SNP) and S-nitroso-N-acetylpenicillamine (SNAP) were used as NO donors. Both minimal and complex media at two pH values, 7.