Time-resolved Förster Resonance Energy Transfer Assays for Measurement of Endogenous Phosphorylated STAT Proteins in Human Cells.

The Janus kinase (JAK)/signal transducer and activator of transcription (STAT) signaling pathway plays a crucial role in mediating cellular responses to cytokines and growth factors. STAT proteins are activated by tyrosine phosphorylation mediated mainly by JAKs. The abnormal activation of STAT signaling pathways is implicated in many human diseases, especially cancer and immune-related conditions.

A sensitive, homogeneous, and high-throughput assay for lysine-specific histone demethylases at the H3K4 site.

Histone methylation is a regulated feature of nucleosomes that can have an impact on gene expression. The methylation state of histone residues has also been found in recent years to be associated with various disorders. Tools for detecting methylation state changes are very useful for dissecting the function of these epigenetic marks.

Development of homogeneous nonradioactive methyltransferase and demethylase assays targeting histone H3 lysine 4.

Histone posttranslational modifications are among the epigenetic mechanisms that modulate chromatin structure and gene transcription. Histone methylation and demethylation are dynamic processes controlled respectively by histone methyltransferases (HMTs) and demethylases (HDMs). Several HMTs and HDMs have been implicated in cancer, inflammation, and diabetes, making them attractive targets for drug therapy.

Development of a high-throughput AlphaScreen assay measuring full-length LRRK2(G2019S) kinase activity using moesin protein substrate.

Mutations within the LRRK2 (leucine-rich repeat kinase 2) gene predispose humans to develop late-onset Parkinson's disease (PD). The most prevalent of these mutations, G2019S, has been shown to increase LRRK2 kinase activity. Therefore, the discovery of small molecule inhibitors of LRRK2(G2019S) through high-throughput screening (HTS) may provide a novel therapeutic strategy for treating PD.

Single-well monitoring of protein-protein interaction and phosphorylation-dephosphorylation events.

We combined oxygen channeling assays with two distinct chemiluminescent beads to detect simultaneously protein phosphorylation and interaction events that are usually monitored separately. This novel method was tested in the ERK1/2 MAP kinase pathway. It was first used to directly monitor dissociation of MAP kinase ERK2 from MEK1 upon phosphorylation and to evaluate MAP kinase phosphatase (MKP) selectivity and mechanism of action.

Metabolic interactions between low doses of benzo[a]pyrene and tributyltin in arctic charr (Salvelinus alpinus): a long-term in vivo study.

We have previously reported that short-term, single exposure to a high dose of tributyltin (TBT), a widely used antifouling biocide, inhibited both the in vivo metabolism and metabolic activation of the carcinogenic polycyclic aromatic hydrocarbon benzo[a]pyrene (BaP) in fish; (BaP), in turn, stimulated TBT metabolism. Here, we provide further mechanistic evidence of mutual metabolic interactions between BaP and TBT in response to long-term, repeated exposures to low doses. Juvenile Arctic charr (Salvelinus alpinus) received 10 separate i.