Autophagy is a lysosome-dependent cellular catabolic mechanism mediating the turnover of intracellular organelles and long-lived proteins. Reduction of autophagy activity has been shown to lead to the accumulation of misfolded proteins in neurons and may be involved in chronic neurodegenerative diseases such as Huntington's disease and Alzheimer's disease. To explore the mechanism of autophagy and identify small molecules that can activate it, we developed a series of high-throughput image-based screens for small-molecule regulators of autophagy. This series of screens allowed us to distinguish compounds that can truly induce autophagic degradation from those that induce the accumulation of autophagosomes as a result of causing cellular damage or blocking downstream lysosomal functions. Our analyses led to the identification of eight compounds that can induce autophagy and promote long-lived protein degradation. Interestingly, seven of eight compounds are FDA-approved drugs for treatment of human diseases. Furthermore, we show that these compounds can reduce the levels of expanded polyglutamine repeats in cultured cells. Our studies suggest the possibility that some of these drugs may be useful for the treatment of Huntington's and other human diseases associated with the accumulation of misfolded proteins.A utophagy is a cellular catabolic mechanism mediating the turnover of intracellular organelles and proteins through a lysosome-dependent but proteasome-independent degradative pathway (1, 2). An autophagosome sequesters cytoplasmic constituents, such as mitochondria, endoplasmic reticulum, and ribosomes, by forming a double-membrane vesicle. The outer membrane of the autophagosome then fuses with the lysosome in mammalian cells delivering the sequestered content to the lumen of lysosome for degradation. Autophagy is critical for the survival of yeast and mammalian cells under starvation conditions because it functions to recycle intracellular material for macromolecular synthesis and energy production (3).Autophagy occurs in all cells at low basal levels under normal conditions to perform homeostatic functions, but it can be rapidly up-regulated under starvation or stress conditions (3). Elegant genetic analysis has identified 17 genes that are essential for autophagy in yeast (referred to as the ATG genes) (4, 5). In mammalian cells, mTOR kinase, the target of rapamycin, mediates the major inhibitory signal that shuts off autophagy under nutrient-rich conditions (3). On the other hand, mammalian type III PI3-kinase, the homolog of yeast VPS34 and inhibitable by 3-methyladenine (3-MA) (a nonspecific inhibitor of PI3-kinase), is required for the onset of autophagy. In this regard, rapamycin and 3-MA, the most commonly used chemicals to induce and inhibit autophagy, respectively, provide convenient tools to study autophagy.To explore the mechanism of autophagy and identify additional small molecules that can activate it, we developed a high-throughput image-based screen. This system takes advantage of the local...

Highwire, a conserved axonal E3 ubiquitin ligase, regulates the initiation of axonal degeneration after injury in Drosophila by regulating the levels of the NAD+ biosynthetic enzyme, Nmnat, and the Wnd kinase.

BackgroundBisphenol A (BPA) is a widespread endocrine-disrupting chemical that can affect humans and animals.ObjectivesWe investigated the effects of adult or prenatal exposure to BPA on T-helper (TH)1/TH2 immune responses and the mechanisms underlying these effects.MethodsTo evaluate the effects of exposure to BPA in adulthood, male Leishmania major–susceptible BALB/c and –resistant C57BL/6 mice were subcutaneously injected with 0.625, 1.25, 2.5, and 5 μmol BPA 1 week before being infected with L. major. To evaluate prenatal exposure, female mice were given BPA-containing drinking water at concentrations of 1, 10, and 100 nM for 2 weeks, then mated, and given BPA for another week. Male 10-week-old offspring were infected with L. major. Footpad swelling was assessed as a measure of the course of infection.ResultsMice exposed to BPA prenatally or in adulthood showed a dose-dependent increase in footpad swelling after being infected with L. major. Exposure to BPA in adulthood significantly promoted antigen-stimulated production of interleukin (IL)-4, IL-10, and IL-13 but not interferon-γ (IFN-γ). However, mice prenatally exposed to BPA showed increased production of not only IL-4 but also IFN-γ. The percentages of CD4+CD25+ cells were decreased in mice exposed to BPA either prenatally or in adulthood. Effects of prenatal BPA exposure were far more pronounced than effects of exposure in adulthood.ConclusionBPA promotes the development of TH2 cells in adulthood and both TH1 and TH2 cells in prenatal stages by reducing the number of regulatory T cells.