The aryl hydrocarbon receptor (Ahr) is a ligand-activated transcription factor that binds DNA in the form of a heterodimer with the Ahr nuclear translocator (hypoxia-inducible factor 1␤). We found in this study that Ahr contains both nuclear localization and export signals in the NH 2 -terminal region. A fusion protein composed of ␤-galactosidase and full-length Ahr translocates from the cytoplasm to the nucleus in a liganddependent manner. However, a fusion protein lacking the PAS (Per-Ahr nuclear translocator-Sim homology) domain of the Ahr showed strong nuclear localization activity irrespective of the presence or absence of ligand. A minimum bipartite Ahr nuclear localization signal (NLS) consisting of amino acid residues 13-39 was identified by microinjection of fused proteins with glutathione S-transferase-green fluorescent protein. A NLS having mutations in bipartite basic amino acids lost nuclear translocation activity completely, which may explain the reduced binding activity to the NLS receptor, PTAC58. A 21-amino acid peptide (residues 55-75) containing the Ahr nuclear export signal is sufficient to direct nuclear export of a microinjected complex of glutathione S-transferase-Ahr-green fluorescent protein. These findings strongly suggest that Ahr act as a ligandand signal-dependent nucleocytoplasmic shuttling protein.The aryl hydrocarbon receptor (Ahr) 1 binds a variety of environmentally important carcinogens, including polycyclic aromatic hydrocarbons and certain halogenated aromatic hydrocarbons such as 2,3,7,8-tetrachlorodibenzo-p-dioxin. Before binding ligands, Ahr is located in the cytoplasm as one component of a complex that has a molecular mass of about 280 kDa (1). This complex is composed of Ahr, two molecules of the 90-kDa heat shock protein, and possibly a 43-kDa protein (2). After ligand binding, Ahr dissociates from the complex and translocates to the nucleus (3). The heterodimer of Ahr and Ahr nuclear translocator (ARNT) constitutes a transcription factor and binds specific DNA sequences called XREs (xenobioticresponsive elements) in the enhancer regions of the CYP1A1 and several other proteins involved in xenobiotic metabolism (4). Because these enzymes are involved in the metabolism of polycyclic aromatic hydrocarbons to active genotoxic metabolites, Ahr plays an important role in carcinogenesis caused by these compounds (5-7).Because ARNT was first cloned as a factor required for ligand-dependent nuclear translocation of Ahr from the cytoplasm to the nucleus (8), the subcellular localization of ARNT was believed to be cytoplasmic. In fact, most ARNT was recovered in the cytosolic fraction by cell fractionation. However, immunohistochemical analysis has shown that ARNT is localized predominantly in the nucleus, regardless of the presence or absence of ligands (9, 10). This controversial subject was clarified by our recent study in which a nuclear localization signal (NLS) of the amino acid residues between 39 and 61 of human ARNT was found to be a novel bipartite type recognized by th...

Intestinal cancer is one of the most common human cancers. Aberrant activation of the canonical Wnt signaling cascade, for example, caused by adenomatous polyposis coli (APC) gene mutations, leads to increased stabilization and accumulation of ␤-catenin, resulting in initiation of intestinal carcinogenesis. The aryl hydrocarbon receptor (AhR) has dual roles in regulating intracellular protein levels both as a ligand-activated transcription factor and as a ligand-dependent E3 ubiquitin ligase. Here, we show that the AhR E3 ubiquitin ligase has a role in suppression of intestinal carcinogenesis by a previously undescribed ligand-dependent ␤-catenin degradation pathway that is independent of and parallel to the APC system. This function of AhR is activated by both xenobiotics and natural AhR ligands, such as indole derivatives that are converted from dietary tryptophan and glucosinolates by intestinal microbes, and suppresses intestinal tumor development in Apc Min/؉ mice. These findings suggest that chemoprevention with naturally-occurring and chemically-designed AhR ligands can be used to successfully prevent intestinal cancers.cecal cancer ͉ ubiquitin ligase ͉ ␤-catenin ͉ tumor chemoprevention

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Cytochrome P450 1B1 (CYP1B1) participates in the metabolic activation of a number of procarcinogens including benzo[a]pyrene and the hydroxylation of 17beta-estradiol at the C-4 position. In this study, we investigated the association between CYP1B1 genetic polymorphism and breast or lung cancer incidence. The Ala-Ser polymorphism at codon 119 in presumed substrate recognition site 1 was significantly associated with the incidence of breast or squamous cell carcinoma of the lung. On the other hand, Leu-Val polymorphism at codon 432 did not show any association to the cancers. An allele containing both Ala and Leu simultaneously, comprised 75% of alleles among 315 Japanese healthy controls, was significantly inversely associated with breast cancer incidence. When expressed in a recombinant system, this CYP1B1 cDNA showed the lowest 17beta-estradiol 4-hydroxylase activity among four different variant forms of CYP1B1. Thus, inter-individual differences in activation of procarcinogens or metabolism of oestrogen originating from genetic polymorphisms of the human CYP1B1 gene may contribute to the susceptibility of human cancers.

Ku antigen is a complex of Ku70 and Ku80 subunits and plays an important role in not only DNA double-strand breaks (DSB) repair and V(D)J recombination, but also in growth regulation. Ku is generally believed to always form and function as heterodimers on the basis of in vitro observations. Here we demonstrate that the localization of Ku80 does not completely coincide with that of Ku70. Ku70 and Ku80 were colocalized in the nucleus in the interphase but not in the late telophase/early G1 phase of the cell cycle. Since the in vivo function of Ku might be partially regulated by the control of its transport, we attempted to investigate the molecular mechanisms underlying the nuclear translocation of Ku. The nuclear translocation of Ku80 started during the late telophase/ early G1 phase after the nuclear envelope was formed and this was preceded by the nuclear translocation of Ku70. Furthermore, we found that the Ku80 protein was transported to the nucleus without heterodimerization with Ku70. To understand in detail the mechanism of transport of Ku80, we attempted to identify the nuclear localization signal (NLS) of Ku80 and de®ned to a region spanning nine amino acid residues (positions 561 ± 569). The Ku80 NLS was demonstrated to be mediated to the nuclear rim by two components of PTAC58 and PTAC97. All these ®ndings support the idea that Ku80 can translocate to the nucleus using its own NLS independent of the translocation of Ku70.