Robustness of NanoBiT luciferase complementation technology in the presence of widely used kinase inhibitors.

Bioluminescence assays using luciferase enzymes are widely used in research to monitor gene expression and an array of other cell properties, and split luciferase enzymes can be used to measure protein interactions in biochemical assays and in living cells. When these methods are employed in chemical library screening efforts, it is vital that the activity of the luciferase enzyme itself is not strongly influenced by library components. Here, we developed a NanoBiT split luciferase assay to measure phosphorylation of Histone H3 peptides and used it to test the robustness of split luciferase to interference from two libraries of commonly used kinase inhibitors, including the Kinase Chemogenomic Set (KCGS).

Dissecting the roles of Haspin and VRK1 in histone H3 phosphorylation during mitosis.

Protein kinases that phosphorylate histones are ideally-placed to influence the behavior of chromosomes during cell division. Indeed, a number of conserved histone phosphorylation events occur prominently during mitosis and meiosis in most eukaryotes, including on histone H3 at threonine-3 (H3T3ph). At least two kinases, Haspin and VRK1 (NHK-1/ballchen in Drosophila), have been proposed to carry out this modification.

The toxicity of the methylimidazolium ionic liquids, with a focus on M8OI and hepatic effects.

Ionic liquids are a diverse range of charged chemicals with low volatility and often liquids at ambient temperatures. This characteristic has in part lead to them being considered environmentally-friendly replacements for existing volatile solvents. However, methylimidazolium ionic liquids are slow to break down in the environment and a recent study at Newcastle detected 1 octyl 3 methylimidazolium (M8OI) - an 8 carbon variant methylimidazolium ionic liquid - in soils in close proximity to a landfill site.

Phosphorylation of MCPH1 isoforms during mitosis followed by isoform-specific degradation by APC/C-CDH1.

Microcephalin-1 (MCPH1) exists as 2 isoforms that regulate cyclin-dependent kinase-1 activation and chromosome condensation during mitosis, with MCPH1 mutations causing primary microcephaly. MCPH1 is also a tumor suppressor protein, with roles in DNA damage repair/checkpoints. Despite these important roles, there is little information on the cellular regulation of MCPH1.

Identification of a xenobiotic as a potential environmental trigger in primary biliary cholangitis.

Background & Aims: Primary biliary cholangitis (PBC) is an autoimmune-associated chronic liver disease triggered by environmental factors, such as exposure to xenobiotics, which leads to a loss of tolerance to the lipoic acid-conjugated regions of the mitochondrial pyruvate dehydrogenase complex, typically to the E2 component. We aimed to identify xenobiotics that might be involved in the environmental triggering of PBC.

Methods: Urban landfill and control soil samples from a region with high PBC incidence were screened for xenobiotic activities using analytical, cell-based xenobiotic receptor activation assays and toxicity screens.

B-13 progenitor-derived hepatocytes (B-13/H cells) model lipid dysregulation in response to drugs and chemicals.

Lipid dysregulation is a common hepatic adverse outcome after exposure to toxic drugs and chemicals. A donor-free rat hepatocyte-like (B-13/H) cell was therefore examined as an in vitro model for investigating mechanisms. The B-13/H cell irreversibly accumulated triglycerides (steatosis) in a time- and dose-dependent manner when exposed to fatty acids, an effect that was potentiated by the combined addition of hyperglycaemic levels of glucose and insulin.

Hepatic effects of tartrazine (E 102) after systemic exposure are independent of oestrogen receptor interactions in the mouse.

Tartrazine is a food colour that activates the transcriptional function of the human oestrogen receptor alpha in an in vitro cell model. Since oestrogens are cholestatic, we hypothesised tartrazine will cause periportal injury to the liver in vivo. To test this hypothesis, tartrazine was initially administered systemically to mice resulting in a periportal recruitment of inflammatory cells, increased serum alkaline phosphatase activity and mild periportal fibrosis.

Environmental Xenoestrogens Super-Activate a Variant Murine ER Beta in Cholangiocytes.

High systemic levels of oestrogens are cholestatic and primary biliary cholangitis (PBC)-which is characterized by hepatic ductular inflammation-is thought to be triggered by exposure to xenobiotics such as those around landfill sites. Xenoestrogens may be a component of this chemical trigger. We therefore hypothesized that xenoestrogens are present at higher levels in the proximity of landfill sites.