HIV-1 Tat Promotes Kaposi's Sarcoma-Associated Herpesvirus (KSHV) vIL-6-Induced Angiogenesis and Tumorigenesis by Regulating PI3K/PTEN/AKT/GSK-3β Signaling Pathway

Kaposi's sarcoma (KS)-associated herpesvirus (KSHV) is etiologically associated with KS, the most common AIDS-related malignancy. KS is characterized by vast angiogenesis and hyperproliferative spindle cells. We have previously reported that HIV-1 Tat can trigger KSHV reactivation and accelerate Kaposin A-induced tumorigenesis. Here, we explored Tat promotion of KSHV vIL-6-induced angiogenesis and tumorigenesis. Tat promotes vIL-6-induced cell proliferation, cellular transformation, vascular tube formation and VEGF production in culture. Tat enhances vIL-6-induced angiogenesis and tumorigenesis of fibroblasts and human endothelial cells in a chicken chorioallantoic membrane (CAM) model. In an allograft model, Tat promotes vIL-6-induced tumorigenesis and expression of CD31, CD34, SMA, VEGF, b-FGF, and cyclin D1. Mechanistic studies indicated Tat activates PI3K and AKT, and inactivates PTEN and GSK-3β in vIL-6 expressing cells. LY294002, a specific inhibitor of PI3K, effectively impaired Tat's promotion of vIL-6-induced tumorigenesis. Together, these results provide the first evidence that Tat might contribute to KS pathogenesis by synergizing with vIL-6, and identify PI3K/AKT pathway as a potential therapeutic target in AIDS-related KS patients. © 2013 Zhou et al

Primary DNA damage and genetic polymorphisms for CYP1A1, EPHX and GSTM1 in workers at a graphite electrode manufacturing plant

<p>Abstract</p> <p>Background</p> <p>The results of a cross-sectional study aimed to evaluate whether genetic polymorphisms (biomarkers of susceptibility) for <it>CYP1A1</it>, <it>EPHX </it>and <it>GSTM1 </it>genes that affect polycyclic aromatic hydrocarbons (PAH) activation and detoxification might influence the extent of primary DNA damage (biomarker of biologically effective dose) in PAH exposed workers are presented. PAH-exposure of the study populations was assessed by determining the concentration of 1-hydroxypyrene (1OHP) in urine samples (biomarker of exposure dose).</p> <p>Methods</p> <p>The exposed group consisted of workers (n = 109) at a graphite electrode manufacturing plant, occupationally exposed to PAH. Urinary 1OHP was measured by HPLC. Primary DNA damage was evaluated by the alkaline comet assay in peripheral blood leukocytes. Genetic polymorphisms for <it>CYP1A1</it>, <it>EPHX</it> and <it>GSTM1</it> were determined by PCR or PCR/RFLP analysis.</p> <p>Results</p> <p>1OHP and primary DNA damage were significantly higher in electrode workers compared to reference subjects. Moreover, categorization of subjects as normal or outlier highlighted an increased genotoxic risk OR = 2.59 (CI95% 1.32–5.05) associated to exposure to PAH. Polymorphisms in <it>EPHX</it> exons 3 and 4 was associated to higher urinary concentrations of 1OHP, whereas none of the genotypes analyzed (<it>CYP1A1</it>, <it>EPHX</it>, and <it>GSTM1</it>) had any significant influence on primary DNA damage as evaluated by the comet assay.</p> <p>Conclusion</p> <p>The outcomes of the present study show that molecular epidemiology approaches (i.e. cross-sectional studies of genotoxicity biomarkers) can play a role in identifying common genetic risk factors, also attempting to associate the effects with measured exposure data. Moreover, categorization of subjects as normal or outlier allowed the evaluation of the association between occupational exposure to PAH and DNA damage highlighting an increased genotoxic risk.</p

Domain Structure of the NRIF3 Family of Coregulators Suggests Potential Dual Roles in Transcriptional Regulation

The identification of a novel coregulator for nuclear hormone receptors, designated NRIF3, was recently reported (D. Li et al., Mol. Cell. Biol. 19:7191–7202, 1999). Unlike most known coactivators, NRIF3 exhibits a distinct receptor specificity in interacting with and potentiating the activity of only TRs and RXRs but not other examined nuclear receptors. However, the molecular basis underlying such specificity is unclear. In this report, we extended our study of NRIF3-receptor interactions. Our results suggest a bivalent interaction model, where a single NRIF3 molecule utilizes both the C-terminal LXXIL (receptor-interacting domain 1 [RID1]) and the N-terminal LXXLL (RID2) modules to cooperatively interact with TR or RXR (presumably a receptor dimer), with the spacing between RID1 and RID2 playing an important role in influencing the affinity of the interactions. During the course of these studies, we also uncovered an NRIF3-NRIF3 interaction domain. Deletion and mutagenesis analyses mapped the dimerization domain to a region in the middle of NRIF3 (residues 84 to 112), which is predicted to form a coiled-coil structure and contains a putative leucine zipper-like motif. By using Gal4 fusion constructs, we identified an autonomous transactivation domain (AD1) at the C terminus of NRIF3. Somewhat surprisingly, full-length NRIF3 fused to the DNA-binding domain of Gal4 was found to repress transcription of a Gal4 reporter. Further analyses mapped a novel repression domain (RepD1) to a small region at the N-terminal portion of NRIF3 (residues 20 to 50). The NRIF3 gene encodes at least two additional isoforms due to alternative splicing. These two isoforms contain the same RepD1 region as NRIF3. Consistent with this, Gal4 fusions of these two isoforms were also found to repress transcription. Cotransfection of NRIF3 or its two isoforms did not relieve the transrepression function mediated by their corresponding Gal4 fusion proteins, suggesting that the repression involves a mechanism(s) other than the recruitment of a titratable corepressor. Interestingly, a single amino acid residue change of a potential phosphorylation site in RepD1 (Ser(28) to Ala) abolishes its transrepression function, suggesting that the coregulatory property of NRIF3 (or its isoforms) might be subjected to regulation by cellular signaling. Taken together, our results identify NRIF3 as an interesting coregulator that possesses both transactivation and transrepression domains and/or functions. Collectively, the NRIF3 family of coregulators (which includes NRIF3 and its other isoforms) may play dual roles in mediating both positive and negative regulatory effects on gene expression

Activation of NF-κB by the Human Herpesvirus 8 Chemokine Receptor ORF74: Evidence for a Paracrine Model of Kaposi's Sarcoma Pathogenesis

Infection with human herpesvirus 8 (HHV-8), also known as Kaposi's sarcoma (KS)-associated herpesvirus, is necessary for the development of KS. The HHV-8 lytic-phase gene ORF74 is related to G protein-coupled receptors, particularly interleukin-8 (IL-8) receptors. ORF74 activates the inositol phosphate/phospholipase C pathway and the downstream mitogen-activated protein kinases, JNK/SAPK and p38. We show here that ORF74 also activates NF-κB independent of ligand when expressed in KS-derived HHV-8-negative endothelial cells or primary vascular endothelial cells. NF-κB activation was enhanced by the chemokine GROα, but not by IL-8. Mutation of Val to Asp in the ORF74 second cytoplasmic loop did not affect ligand-independent signaling activity, but it greatly increased the response to GROα. ORF74 upregulated the expression of NF-κB-dependent inflammatory cytokines (RANTES, IL-6, IL-8, and granulocyte-macrophage colony-stimulating factor) and adhesion molecules (VCAM-1, ICAM-1, and E-selectin). Supernatants from transfected KS cells activated NF-κB signaling in untransfected cells and elicited the chemotaxis of monocytoid and T-lymphoid cells. Expression of ORF74 conferred on primary endothelial cells a morphology that was strikingly similar to that of spindle cells present in KS lesions. Taken together, these data, demonstrating that ORF74 activates NF-κB and induces the expression of proangiogenic and proinflammatory factors, suggest that expression of ORF74 in a minority of cells in KS lesions could influence uninfected cells or latently infected cells via autocrine and paracrine mechanisms, thereby contributing to KS pathogenesis