Increases in the intracellular levels of reactive oxygen species (ROS), frequently referred to as oxidative stress, represents a potentially toxic insult which if not counteracted will lead to membrane dysfunction, DNA damage and inactivation of proteins. Chronic oxidative stress has numerous pathological consequences including cancer, arthritis and neurodegenerative disease. Glutathione-associated metabolism is a major mechanism for cellular protection against agents which generate oxidative stress. It is becoming increasingly apparent that the glutathione tripeptide is central to a complex multifaceted detoxification system, where there is substantial inter-dependence between separate component members. Glutathione participates in detoxification at several different levels, and may scavenge free radicals, reduce peroxides or be conjugated with electrophilic compounds. Thus, glutathione provides the cell with multiple defences not only against ROS but also against their toxic products. This article discusses how glutathione biosynthesis, glutathione peroxidases, glutathione S-transferases and glutathione S-conjugate efflux pumps function in an integrated fashion to allow cellular adaption to oxidative stress. Co-ordination of this response is achieved, at least in part, through the antioxidant responsive element (ARE) which is found in the promoters of many of the genes that are inducible by oxidative and chemical stress. Transcriptional activation through this enhancer appears to be mediated by basic leucine zipper transcription factors such as Nrf and small Maf proteins. The nature of the intracellular sensor(s) for ROS and thiol-active chemicals which induce genes through the ARE is described. Gene activation through the ARE appears to account for the enhanced antioxidant and detoxification capacity of normal cells effected by many cancer chemopreventive agents. In certain instances it may also account for acquired resistance of tumours to cancer chemotherapeutic drugs. It is therefore clear that determining the mechanisms involved in regulation of ARE-driven gene expression has enormous medical implications.

Glutathione and glutathione-dependent enzymes represent a co-ordinately regulated defence against oxidative stress

In this article, the history of the LDL receptor is recounted by its codiscoverers. Their early work on the LDL receptor explained a genetic cause of heart attacks and led to new ways of thinking about cholesterol metabolism. The LDL receptor discovery also introduced three general concepts to cell biology: receptor-mediated endocytosis, receptor recycling, and feedback regulation of receptors. The latter concept provides the mechanism by which statins selectively lower plasma LDL, reducing heart attacks and prolonging life.

The LDL Receptor

Niemann-Pick disease type C (NPC) is an autosomal recessive neurovisceral lipid storage with a wide spectrum of clinical phenotypes. At the cellular level, the disorder is characterized by accumulation of unesterified cholesterol and glycolipids in the lysosomal/late endosomal system. Approximatively 95% of patients have mutations in the NPC1 gene (mapped at 18q11) which encodes a large membrane glycoprotein primarily located to late endosomes. The remainder have mutations in the NPC2 gene (mapped at 14q24.3) which encodes a small soluble lysosomal protein with cholesterol-binding properties. The identical biochemical patterns observed in NPC1 and NPC2 mutants suggest that the two proteins function in a coordinate fashion. Identification of mutations revealed a complex picture of molecular heterogeneity, allowing genotype - phenotype correlations for both genes and providing insights into structure - function relationships for the NPC1 protein. Although a whole body of evidence suggests that the NPC1 and NPC2 proteins are involved in the cellular postlysosomal/late endosomal transport of cholesterol, glycolipids and other cargo, their precise functions and relationship remain unclear and are currently the subject of intense investigation. These studies, conducted in various models, should ultimately lead to a better understanding of the pathophysiology of NPC and new therapeutic approaches.

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Niemann-Pick disease type C.

Structure of N-terminal domain of NPC1 reveals distinct subdomains for binding and transfer of cholesterol.

The steroidogenic acute regulatory protein (StAR) regulates acute steroidogenesis in the adrenal cortex and gonads by promoting the translocation of cholesterol to the mitochondrial inner membrane where the first step in steriod biosynthesis is catalyzed. StAR-related lipid transfer (START) domains occur in proteins involved in lipid transport and metabolism, signal transduction, and transcriptional regulation. The 2.2 A resolution crystal structure of the START domain of human MLN64 reported here reveals an alpha/beta fold built around a U-shaped incomplete beta-barrel. The interior of the protein encompasses a 26 x 12 x 11 A hydrophobic tunnel that is large enough to bind a single cholesterol molecule. The StAR and MLN64 START domains bind 1 mole of 14C cholesterol per mole of protein in vitro. Based on the START domain structure and cholesterol binding stoichiometry, it is proposed that StAR acts by shuttling cholesterol molecules one at a time through the intermembrane space of the mitochondrion.

Shinsuke Hiramoto

Papers

A porcine homolog of the major secretory protein of human epididymis, HE1, specifically binds cholesterol.

Molecular cloning and characterization of the epididymis-specific glutathione peroxidase-like protein secreted in the porcine epididymal fluid.

Stage-specific expression of a mouse homologue of the porcine 135kDa alpha-D-mannosidase (MAN2B2) in type A spermatogonia.