Real Time Fluorescence Imaging of Plcγ Translocation and Its Interaction with the Epidermal Growth Factor Receptor

The translocation of fluorescently tagged PLCγ and requirements for this process in cells stimulated with EGF were analyzed using real time fluorescence microscopy applied for the first time to monitor growth factor receptor–effector interactions. The translocation of PLCγ to the plasma membrane required the functional Src homology 2 domains and was not affected by mutations in the pleckstrin homology domain or inhibition of phosphatidylinositol (PI) 3-kinase. An array of domains specific for PLCγ isoforms was sufficient for this translocation. The dynamics of translocation to the plasma membrane and redistribution of PLCγ, relative to localization of the EGF receptor and PI 4,5-biphosphate (PI 4,5-P2), were shown. Colocalization with the receptor was observed in the plasma membrane and in membrane ruffles where PI 4,5-P2 substrate could also be visualized. At later times, internalization of PLCγ, which could lead to separation from the substrate, was observed. The data support a direct binding of PLCγ to the receptor as the main site of the plasma membrane recruitment. The presence of PLCγ in membrane structures and its access to the substrate appear to be transient and are followed by a rapid incorporation into intracellular vesicles, leading to downregulation of the PLC activity.</jats:p

Applications of microarray technology in breast cancer research

Microarrays provide a versatile platform for utilizing information from the Human Genome Project to benefit human health. This article reviews the ways in which microarray technology may be used in breast cancer research. its diverse applications include monitoring chromosome gains and losses, tumour classification, drug discovery and development, DNA resequencing, mutation detection and investigating the mechanism of tumour development

Short tandem repeat profiling provides an international reference standard for human cell lines

Cross-contamination between cell lines is a longstanding and frequent cause of scientific misrepresentation. Estimates from national testing services indicate that up to 36% of cell lines are of a different origin or species to that claimed. To test a standard method of cell line authentication, 253 human cell lines from banks and research institutes worldwide were analyzed by short tandem repeat profiling. The short tandem repeat profile is a simple numerical code that is reproducible between laboratories, is inexpensive, and can provide an international reference standard for every cell line. If DNA profiling of cell lines is accepted and demanded internationally, scientific misrepresentation because of cross-contamination can be largely eliminated.</jats:p

A mutation in the Gsk3-binding domain of zebrafish Masterblind/Axin1 leads to a fate transformation of telencephalon and eyes to diencephalon

Zebrafish embryos homozygous for the masterblind (mb1) mutation exhibit a striking phenotype in which the eyes and telencephalon are reduced or absent and diencephalic fates expand to the front of the brain. Here we show that mb1(-/-) embryos carry an amino-acid change at a conserved site in the Wnt pathway scaffolding protein, Axin1. The amino-acid substitution present in the mbl allele abolishes the binding of Axin to Gsk3 and affects Tcf-dependent transcription. Therefore, Gsk3 activity may be decreased in mbl(-/-) embryos and in support of this possibility, overexpression of either wild-type Axin1 or Gsk3 beta can restore eye and telencephalic fates to mb1(-/-) embryos. Our data reveal a crucial role for Axin1-dependent inhibition of the Wnt pathway in the early regional subdivision of the anterior neural plate into telencephalic, diencephalic, and eye-forming territories