Chromophore-assisted laser inactivation--towards a spatiotemporal-functional analysis of proteins, and the ablation of chromatin, organelle and cell function.

Chromophore-assisted laser or light inactivation (CALI) has been employed as a promising technique to achieve spatiotemporal knockdown or loss-of-function of target molecules in situ. CALI is performed using photosensitizers as generators of reactive oxygen species (ROS). There are two CALI approaches that use either transgenic tags with chemical photosensitizers, or genetically encoded fluorescent protein fusions. Using spatially restricted microscopy illumination, CALI can address questions regarding, for example, protein isoforms, subcellular localization or phase-specific analyses of multifunctional proteins that other knockdown approaches, such as RNA interference or treatment with chemicals, cannot. Furthermore, rescue experiments can clarify the phenotypic capabilities of CALI after the depletion of endogenous targets. CALI can also provide information about individual events that are involved in the function of a target protein and highlight them in multifactorial events. Beyond functional analysis of proteins, CALI of nuclear proteins can be performed to induce cell cycle arrest, chromatin- or locus-specific DNA damage. Even at organelle level - such as in mitochondria, the plasma membrane or lysosomes - CALI can trigger cell death. Moreover, CALI has emerged as an optogenetic tool to switch off signaling pathways, including the optical depletion of individual neurons. In this Commentary, we review recent applications of CALI and discuss the utility and effective use of CALI to address open questions in cell biology.

Effects of green tea on gene expression of hepatic gluconeogenic enzymes in vivo.

It has recently been reported that the major green tea polyphenolic constituent, epigallocatechin 3-gallate (EGCG), mimics the cellular effects of insulin including the reductive effect on the gene expression of rate-limiting gluconeogenic enzymes in a cell culture system. We show that administration of green tea that contains EGCG caused a reduction in the level of mRNAs for gluconeogenic enzymes, phosphoenolpyruvate carboxykinase and glucose-6-phosphatase in the mouse liver. EGCG alone was also found to down-regulate the gene expression of these enzymes but not so curcumin or quercetin. The results of this study support the idea that green tea intake may be beneficial in the prevention of diabetes mellitus.