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Apurinic͞apyrimidinic (AP) endonuclease (APE; EC 4.2.99.18) plays a central role in repair of DNA damage due to reactive oxygen species (ROS) because its DNA 3-phosphoesterase activity removes 3 blocking groups in DNA that are generated by DNA glycosylase͞AP-lyases during removal of oxidized bases and by direct ROS reaction with DNA. The major human APE (APE-1) gene is activated selectively by sublethal levels of a variety of ROS and ROS generators, including ionizing radiation, but not by other genotoxicants-e.g., UV light and alkylating agents. Increased expression of APE mRNA and protein was observed both in the HeLa S3 tumor line and in WI 38 primary fibroblasts, and it was accompanied by translocation of the endonuclease to the nucleus. ROS-treated cells showed a significant increase in resistance to the cytotoxicity of such ROS generators as H 2 O 2 and bleomycin, but not to UV light. This ''adaptive response'' appears to result from enhanced repair of cytotoxic DNA lesions due to an increased activity of APE-1, which may be limiting in the base excision repair process for ROSinduced toxic lesions.Reactive oxygen species (ROS), including partially reduced oxygen species, namely, superoxide anion (O 2 ⅐ -), H 2 O 2 , and the hydroxyl radical ( ⅐ OH), are generated endogenously as byproducts of respiration and also during pathological states in activated neutrophils (1-3). ROS can also be generated by external agents, such as ionizing radiation (4) and longwavelength UV light (5) and during binding of transition metals and other ligands such as tumor necrosis factor ␣ and interleukin 1 (6). Because of their ubiquitous and continuous presence, these reactive radicals and oxidizing agents have been implicated in the etiology of a variety of diseases, including arthritis, cancer, and neurodegenerative diseases, and also in aging (7-9). While ROS react with most cellular components, they are also invariably genotoxic, because they react with both the deoxyribose and bases in DNA, and so generate a plethora of base lesions and strand breaks (4, 10, 11). Many of these lesions are cytotoxic and mutagenic (12, 13). Most, if not all, of the lesions induced by ROS are repaired via the base excision repair (BER) pathway, in which the damaged bases are removed by specific DNA glycosylases, leaving abasic sites that are subsequently cleaved by apurinic͞ apyrimidinic (AP) endonucleases [APEs; EC 4.2.99.18 (14, 15)]. The major mammalian APE (called APE-1) contributes more than 80% of the total APE activity and is analogous to Escherichia coli exonuclease III (Xth protein) (16)(17)(18)(19). The majority, if not all, of the ROS-altered base lesions in DNA are removed by two DNA glycosylases-endonuclease III (NTH) and 8-oxoguanine-DNA glycosylase (OGG)-in both E. coli and mammalian cells (20)(21)(22)(23)(24)(25). These enzymes have a second activity, AP-lyase, which in a concerted reaction cleaves the base N-glycosyl bond, and carries out ␤ or ␤␦ elimination of the resulting free deoxyribose. They thus cleave the phos...

Stat1 is a fascinating and complex protein with multiple, yet contrasting transcriptional functions. Upon activation, it drives the expression of many genes but also suppresses the transcription of others. These opposing characteristics also apply to its role in facilitating crosstalk between signal transduction pathways, as it participates in both synergistic activation and inhibition of gene expression. Stat1 is a functional transcription factor even in the absence of inducer-mediated activation, participating in the constitutive expression of some genes. This review summarizes the well studied involvement of Stat1 in IFN-dependent and growth factor-dependent signaling and then describes the roles of Stat1 in positive, negative and constitutive regulation of gene expression as well as its participation in crosstalk between signal transduction pathways. Oncogene (2000).

Interferons (IFNs) inhibit cell growth in a Stat1-dependent fashion that involves regulation of c-myc expression. IFN-gamma suppresses c-myc in wild-type mouse embryo fibroblasts, but not in Stat1-null cells, where IFNs induce c-myc mRNA rapidly and transiently, thus revealing a novel signaling pathway. Both tyrosine and serine phosphorylation of Stat1 are required for suppression. Induced expression of c-myc is likely to contribute to the proliferation of Stat1-null cells in response to IFNs. IFNs also suppress platelet-derived growth factor (PDGF)-induced c-myc expression in wild-type but not in Stat1-null cells. A gamma-activated sequence element in the promoter is necessary but not sufficient to suppress c-myc expression in wild-type cells. In PKR-null cells, the phosphorylation of Stat1 on Ser727 and transactivation are both defective, and c-myc mRNA is induced, not suppressed, in response to IFN-gamma. A role for Raf-1 in the Stat1-independent pathway is revealed by studies with geldanamycin, an HSP90-specific inhibitor, and by expression of a mutant of p50(cdc37) that is unable to recruit HSP90 to the Raf-1 complex. Both agents abrogated the IFN-gamma-dependent induction of c-myc expression in Stat1-null cells.

Full activation of STAT11 by IFN␥ requires two distinct phosphorylation events. Receptor-mediated phosphorylation of STAT1 on tyrosine 701 is required for STAT1 homodimers to form and subsequently to bind to the promoters of IFN␥-responsive genes through ␥-activated sequence (GAS) elements (reviewed in Refs. 1 and 2). The mechanistic basis for STAT1 tyrosine phosphorylation has been established for some time. JAK1 and JAK2, constitutively bound to specific cytoplasmic domains of the IFNGR1 and IFNGR2 subunits, phosphorylate each other when IFN␥ binds and the receptor subunits aggregate. The activated JAKs then phosphorylate tyrosine 440 of IFNGR1, creating a docking site that recruits STAT1 to the receptor. STAT1 binds to phosphorylated Tyr-440 through its SH2 domain, allowing its Tyr-701 residue to be phosphorylated by the JAKs. STAT1 then dissociates from the receptor, dimerizes through reciprocal SH2-phosphotyrosine interactions, and binds to IFN␥-inducible promoters.

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PURPOSE More than 80% of patients who undergo sentinel lymph node (SLN) biopsy have no nodal metastasis. Here, we describe a model that combines clinicopathologic and molecular variables to identify patients with thin- and intermediate-thickness melanomas who may forgo the SLN biopsy procedure because of their low risk of nodal metastasis. PATIENTS AND METHODS Genes with functional roles in melanoma metastasis were discovered by analysis of next-generation sequencing data and case-control studies. We then used polymerase chain reaction to quantify gene expression in diagnostic biopsy tissue across a prospectively designed archival cohort of 754 consecutive thin- and intermediate-thickness primary cutaneous melanomas. Outcome of interest was SLN biopsy metastasis within 90 days of melanoma diagnosis. A penalized maximum likelihood estimation algorithm was used to train logistic regression models in a repeated cross-validation scheme to predict the presence of SLN metastasis from molecular, clinical, and histologic variables. RESULTS Expression of genes with roles in epithelial-to-mesenchymal transition (glia-derived nexin, growth differentiation factor 15, integrin-β3, interleukin 8, lysyl oxidase homolog 4, transforming growth factor-β receptor type 1, and tissue-type plasminogen activator) and melanosome function (melanoma antigen recognized by T cells 1) were associated with SLN metastasis. The predictive ability of a model that only considered clinicopathologic or gene expression variables was outperformed by a model that included molecular variables in combination with the clinicopathologic predictors Breslow thickness and patient age (area under the receiver operating characteristic curve, 0.82; 95% CI, 0.78 to 0.86; SLN biopsy reduction rate, 42%; negative predictive value, 96%). CONCLUSION A combined model that included clinicopathologic and gene expression variables improved the identification of patients with melanoma who may forgo the SLN biopsy procedure because of their low risk of nodal metastasis.