The androgen-signaling pathway is important for the growth and progression of prostate cancer cells. The growth-promoting effects of androgen on prostate cells are mediated mostly through the androgen receptor (AR). There is increasing evidence that transcription activation by AR is mediated through interaction with other cofactors. ␤-Catenin plays a critical role in embryonic development and tumorigenesis through its effects on E-cadherin-mediated cell adhesion and Wnt-dependent signal transduction. Here, we demonstrate that a specific protein-protein interaction occurs between ␤-catenin and AR. Unlike the steroid hormone receptor coactivator 1 (SRC1), ␤-catenin showed a strong interaction with AR but not with other steroid hormone receptors such as estrogen receptor ␣, progesterone receptor ␤, and glucocorticoid receptor. The ligand binding domain of AR and the NH 2 terminus combined with the first six armadillo repeats of ␤-catenin were shown to be necessary for the interaction. Through this specific interaction, ␤-catenin augments the ligand-dependent activity of AR in prostate cancer cells. Moreover, expression of E-cadherin in E-cadherin-negative prostate cancer cells results in redistribution of the cytoplasmic ␤-catenin to the cell membrane and reduction of ARmediated transcription. These data suggest that loss of E-cadherin can elevate the cellular levels of ␤-catenin in prostate cancer cells, which may directly contribute to invasiveness and a more malignant tumor phenotype by augmenting AR activity during prostate cancer progression.

The association of the NF-B p65/p50 dimer with IB␣ plays a pivotal role in regulating its nuclear translocation and gene transcription. In addition, serine phosphorylation at various sites of the p65 subunit has been shown to be important in initiating transcription. Here we demonstrate that the regulation of nuclear translocation of p65 phosphorylated at serine 536 is not dependent on IB␣. Stimulation of either Jurkat or normal human T cells resulted in the nuclear translocation of phospho-p65 (Ser 536 ). In addition, the phospho-p65 (Ser 536 ) was not associated with either IB␣ or p50, and the nuclear translocation of phospho-p65 (Ser 536 ), but not total p65, was unaffected by the proteosome inhibitor MG-132, which blocks IB protein degradation and prevents p65/p50 dimer nuclear translocation. Accordingly, the co-expression of a dominant negative mutant of IB␣ blocked the transcriptional activity mediated by wild type but not the dominant positive p65 mutant (S536D). Furthermore, the transfection of the S536D form of p65 led to the induction of interleukin-8 transcription following stimulation, whereas the S536A form, which cannot be phosphorylated at this site, did not. Together, the findings suggest that p65 phosphorylated on serine 536 is not associated with or regulated by IB␣, that it has a distinct set of target genes, and that it may represent a noncanonical NF-B pathway that is independent of IB␣ regulation.The NF-B signaling pathway responds rapidly to a wide range of stimuli (1). Activation leads to the translocation of the transcription factors from the cytoplasm to the nucleus. The NF-B transcription factor consists of two subunits of either homo-or heterodimers of RelA/ p65, c-Rel, and p50. The complexes are held in the cytoplasm and prevented from activating transcription by a class of proteins referred to as inhibitors of NF-B or IB proteins. Upon stimulation, the IB proteins are phosphorylated by one of a number of IB kinases (IKK-␣, -␤, and -␥), ubiquitinylated, and degraded, which thereby allows the NF-B complex to translocate into the nucleus (2). However, recent findings have demonstrated the shuttling of the NF-B complex in and out of the nucleus in the absence of stimulation (3, 4). In addition to nuclear translocation of the NF-B complex, several studies have shown that the NF-B proteins are modified post-translationally, and those changes influence transcriptional activity. Examples of activation-induced posttranslational modifications include the acetylation of p65 to facilitate the retention of the NF-B complex in the nucleus (5, 6). In addition, the S-nitrosylation of cysteine 62 of p50 has been shown to affect the NF-B binding to DNA (7, 8).We have previously described the phosphorylation of various Rel proteins following the stimulation of T cells, and the phosphorylation of p50 increased the DNA binding capacity (9). Several studies have demonstrated the phosphorylation of p65 in response to various stimuli (10). Serine 276 of p65 is phosphorylated by protein kinase A during IB d...

Depending on the type of external signals, T cells can initiate multiple intracellular signaling pathways that can be broadly classified into two groups based on their sensitivity to the immunosuppressive drug cyclosporin A (CsA). Interleukin (IL)-12-mediated interferon (IFN)-␥ production by activated T cells has been shown to be CsA-insensitive. In this report, we demonstrate that the IL-12-induced CsA-resistant pathway of IFN-␥ production is sensitive to rapamycin. Rapamycin treatment resulted in the aberrant recruitment of Stat3, Stat4, and phospho-c-Jun to the genomic promoter region resulting in decreased IFN-␥ transcription. IL-12-induced phosphorylation of Stat3 on Ser-727 was affected by rapamycin, which may be due to the effect of rapamycin on the IL-12-induced interaction between mammalian target of rapamycin (mTOR) and Stat3. In accordance with this, reduction in the mTOR protein level by small interfering RNA resulted in suppression of Stat3 phosphorylation and decreased production of IFN-␥ after IL-12 stimulation. These results suggest that mTOR may play a major role in IL-12-induced IFN-␥ production by activated T cells.During an immune reaction in which antigen-presenting cells interact with T cells, both cell types get activated expressing both cell surface molecules and soluble factors that in turn are involved in further activating each cell type. The cell surface molecules CD40 ligand and IL-12-1 and IL-18 receptors, among others, are expressed on T cells, whereas B7 molecules IL-12 and IL-18 are expressed by antigen-presenting cells. One of the cytokines produced by activated T cells is IFN-␥, which is a potent activator of antigen-presenting cells. Activated antigen-presenting cells produce IL-12 and IL-18, which are potent activators of T cells, which in turn produce IFN-␥. This circular pathway of mutual stimulation is important in eliciting a complete cell-mediated immune response.IL-12 is an important cytokine involved in the production of IFN-␥ by T cells and NK cell, and plays an important role in the differentiation of T helper 1 (Th1) cell population (1-4). A unique aspect of IL-12 signaling is that it works together with IL-18 and the T cell co-receptor CD28 in inducing IFN-␥ production (1, 5, 6). Although IL-12-induced IFN-␥ expression has been shown to occur mostly at the level of transcription, the synergy between IL-12 and ␣CD28 or IL-12 and IL-2 has been shown to occur mostly at the post-transcriptional level, e.g. message stability and mobilization of nuclear mRNA (7-9).Upon binding to its receptor, IL-12 activates Janus family kinases Tyk2 and Jak2, which then phosphorylate the tyrosine residues of Stat3 and Stat4 (10). These tyrosine phosphorylations are responsible for the formation of Stat4/Stat4 homodimer and Stat3/Stat4 heterodimers (10 -12). These dimers then translocate to the nucleus and bind to the promoters of IL-12-responsive genes (11, 13). IL-12 stimulation also induces serine phosphorylation of Stat3 and Stat4, which are important for their transcriptional activ...

The mechanisms for "gain-of-function" phenotypes produced by mutant p53s such as enhanced proliferation, resistance to transforming growth factor-␤-mediated growth suppression, and increased tumorigenesis are not known. One theory is that these phenotypes are caused by novel transcriptional regulatory events acquired by mutant p53s. Another explanation is that these effects are a result of an imbalance of functions caused by the retention of some of the wild-type transcriptional regulatory events in the context of a loss of other counterbalancing activities. An analysis of the ability of DNA-binding domain mutants A138P and R175H, and wild-type p53 to regulate the expression levels of 6.9 ؋ 10 3 genes revealed that the mutants retained only <5% of the regulatory activities of the wild-type protein. A138P p53 exhibited mostly retained wild-type regulatory activities and few acquired novel events. However, R175H p53 possessed an approximately equal number of wild-type regulatory events and novel activities. This is the first report that, after examination of the regulation of a large unfocused set of genes, provides data indicating that remaining wild-type transcriptional regulatory functions existing in the absence of counterbalancing activities as well as acquired novel events both contribute to the gain-of-function phenotypes produced by mutant p53s. However, mutant p53s are likely to be distinct in terms of the extent to which each mechanism contributes to their gain-of-function phenotypes.

Objective Multiple sclerosis (MS) is a debilitating neurological disorder involving an autoimmune reaction to oligodendrocytes and degeneration of the axons they ensheath in the CNS. Because the damage to oligodendrocytes and axons involves local inflammation and associated oxidative stress, we tested the therapeutic efficacy of combined treatment with a potent anti-inflammatory thalidomide analog (lenalidomide) and novel synthetic anti-oxidant cerium oxide nanoparticles (nanoceria) in the experimental autoimmune encephalomyelitis (EAE) mouse model of MS. Methods C57BL/6 mice were randomly assigned to a control (no EAE) group, or one of four myelin oligodendrocyte glycoprotein-induced EAE groups: vehicle, lenalidomide, nanoceria, or lenalidomide plus nanoceria. During a 23 day period, clinical EAE symptoms were evaluated daily, and MRI brain scans were performed at 11-13 days and 20-22 days. Histological and biochemical analyses of brain tissue samples were performed to quantify myelin loss and local inflammation. Results Lenalidomide treatment alone delayed symptom onset, while nanoceria treatment had no effect on symptom onset or severity, but did promote recovery; lenalidomide and nanoceria each significantly attenuated white matter pathology and associated inflammation. Combined treatment with lenalidomide and nanoceria resulted in a near elimination of EAE symptoms, and reduced white matter pathology and inflammatory cell responses to a much greater extent than either treatment alone. Interpretation By suppressing inflammation and oxidative stress, combined treatment with lenalidomide and nanoceria can reduce demyelination and associated neurological symptoms in EAE mice. Our preclinical data suggest a potential application of this combination therapy in MS.