Involvement of Rel/Nuclear Factor-κB Transcription Factors in Keratinocyte Senescence

After a finite doubling number, normal cells become senescent, i.e. nonproliferating and apoptosis resistant. Because Rel/nuclear factor (NF)-κB transcription factors regulate both proliferation and apoptosis, we have investigated their involvement in senescence. cRel overexpression in young normal keratinocytes results in premature senescence, as defined by proliferation blockage, apoptosis resistance, enlargement, and appearance of senescence-associated β-galactosidase (SA-β-Gal) activity. Normal senescent keratinocytes display a greater endogenous Rel/NF-κB DNA binding activity than young cells; inhibiting this activity in presenescent cells decreases the number of cells expressing the SA-β-Gal marker. Normal senescent keratinocytes and cRel-induced premature senescent keratinocytes overexpressed manganese superoxide dismutase (MnSOD), a redox enzyme encoded by a Rel/NF-κB target gene. MnSOD transforms the toxic O2.- into H2O2, whereas catalase and glutathione peroxidase convert H2O2 into H2O. Neither catalase nor glutathione peroxidase is up-regulated during cRel-induced premature senescence or during normal senescence, suggesting that H 2O2 accumulates. Quenching H2O2 by catalase delays the occurrence of both normal and premature cRel-induced senescence. Conversely, adding a nontoxic dose of H2O2 to the culture medium of young normal keratinocytes induces a premature senescence-like state. All these results indicate that Rel/NF-κB factors could take part in the occurrence of senescence by generating an oxidative stress via the induction of MnSOD.

The Ets transcription factors of the PEA3 group: Transcriptional regulators in metastasis

The PEA3 group is composed of three highly conserved Ets transcription factors: Erm, Er81, and Pea3. These proteins regulate transcription of multiple genes, and their transactivating potential is affected by post-translational modifications. Among their target genes are several matrix metalloproteases (MMPs), which are enzymes degrading the extracellular matrix during normal remodelling events and cancer metastasis. In fact, PEA3-group genes are often over-expressed in different types of cancers that also over-express these MMPs and display a disseminating phenotype. Experimental regulation of the synthesis of PEA3 group members influences the metastatic process. This suggests that these factors play a key role in metastasis. © 2006 Elsevier B.V. All rights reserved.

Genomic organization of the human e1af gene, a member of Ets transcription factors

The E1AF protein belongs to the family of Ets transcription factors and is involved in the regulation of metastasis gene expression. It has recently been reported in an undifferentiated child sarcoma that part of this gene could be fused by translocation to the ews gene. We show here that the human e1af gene, which is located in the q21 region of chromosome 17, is organized in 13 exons distributed along 19 kb of genomic DNA. Its two main functional domains, the acidic domain and the DNA-binding ETS domain, are each encoded by three different exons. The 3'-untranslated region of e1af is 0.7 kb. The 5'-untranslated region is about 0.3 kb and is composed of a first exon upstream from the exon containing the first methionine. These data could possibly accelerate an understanding of the molecular basis of putative inherited diseases linked to E1AF. (C) 1999 Elsevier Science B.V. All rights reserved.

Differential Modulation of Transcriptional Activity of Estrogen Receptors by Direct Protein-Protein Interactions with the T Cell Factor Family of Transcription Factors

Two major signaling pathways, those triggered by estrogen (E(2)) and by the Wnt family, interact in the breast to cause growth and differentiation. The estrogen receptors ER(alpha) and ER(beta) are activated by binding E(2) and act as ligand-dependent transcription factors. The effector for the Wnt family is the Tcf family of transcription factors. Both sets of transcription factors recognize discrete but different nucleotide sequences in the promoters of their target genes. By using transient transfections of reporter constructs for the osteopontin and thymidine kinase promoters in rat mammary cells, we show that Tcf-4 antagonizes and Tcf-1 stimulates the effects of activated ER/E(2). For mutants of the former promoter, the stimulatory effects of ER(alpha)/E(2) can be made to be dependent on Tcf-1, and for the latter promoter the effects of the T cell factors (TCFs) are dependent on ER/E(2). Direct interaction between ERs and Tcfs either at the Tcf/ER(alpha)-binding site on the DNA or in the absence of DNA is established by gel retardation assays or by coimmunoprecipitation/biosensor methods, respectively. These results show that the two sets of transcription factors can interact directly, the interaction between ERs and Tcf-4 being antagonistic and that between ERs and Tcf-1 being synergistic on the activity of the promoters employed. Since Tcf-4 is the major Tcf family member in the breast, it is suggested that the antagonistic interaction is normally dominant in vivo in this tissue.

The role of pea3 group transcription factors in esophageal squamous cell carcinoma.

The transcription factors Pea3, Erm, and Er81 can promote cancer initiation and progression in various types of solid tumors. However, their role in esophageal squamous cell carcinoma (ESCC) has not been elucidated. In this study, we found that the expression levels of Pea3 and Erm, but not that of Er81, were significantly higher in ESCC compared with nontumor esophageal epithelium. A high level of Pea3 expression was significantly correlated with a shorter overall survival in a cohort of 81 patients with ESCC and the subgroup with N1 stage tumor (Wilcoxon-Gehan test, P = 0.016 and P = 0.001, respectively). Pea3 was overexpressed in seven ESCC cell lines compared with two immortalized esophageal cell lines. Pea3 knockdown reduced cell proliferation and suppressed nonadherent growth, migration, and invasion in ESCC cells in vitro. In addition, Pea3 knockdown in ESCC cells resulted in a down-regulation of phospho-Akt and matrix metalloproteinase 13, whereas a significant positive correlation in the expression levels was observed between Pea3 and phospho-Akt (r = 0.281, P

Pro-neural transcription factors as cancer markers

BACKGROUND: The aberrant transcription in cancer of genes normally associated with embryonic tissue differentiation at various organ sites may be a hallmark of tumour progression. For example, neuroendocrine differentiation is found more commonly in cancers destined to progress, including prostate and lung. We sought to identify proteins which are involved in neuroendocrine differentiation and differentially expressed in aggressive/metastatic tumours.

RESULTS: Expression arrays were used to identify up-regulated transcripts in a neuroendocrine (NE) transgenic mouse model of prostate cancer. Amongst these were several genes normally expressed in neural tissues, including the pro-neural transcription factors Ascl1 and Hes6. Using quantitative RT-PCR and immuno-histochemistry we showed that these same genes were highly expressed in castrate resistant, metastatic LNCaP cell-lines. Finally we performed a meta-analysis on expression array datasets from human clinical material. The expression of these pro-neural transcripts effectively segregates metastatic from localised prostate cancer and benign tissue as well as sub-clustering a variety of other human cancers.

CONCLUSION: By focussing on transcription factors known to drive normal tissue development and comparing expression signatures for normal and malignant mouse tissues we have identified two transcription factors, Ascl1 and Hes6, which appear effective markers for an aggressive phenotype in all prostate models and tissues examined. We suggest that the aberrant initiation of differentiation programs may confer a selective advantage on cells in all contexts and this approach to identify biomarkers therefore has the potential to uncover proteins equally applicable to pre-clinical and clinical cancer biology.

Hydrogen peroxide sensing, signaling and regulation of transcription factors

The regulatory mechanisms by which hydrogen peroxide (H2O2) modulates the activity of transcription factors in bacteria (OxyR and PerR), lower eukaryotes (Yap1, Maf1, Hsf1 and Msn2/4) and mammalian cells (AP-1, NRF2, CREB, HSF1, HIF-1, TP53, NF-κB, NOTCH, SP1 and SCREB-1) are reviewed. The complexity of regulatory networks increases throughout the phylogenetic tree, reaching a high level of complexity in mammalians. Multiple H2O2 sensors and pathways are triggered converging in the regulation of transcription factors at several levels: (1) synthesis of the transcription factor by upregulating transcription or increasing both mRNA stability and translation; (ii) stability of the transcription factor by decreasing its association with the ubiquitin E3 ligase complex or by inhibiting this complex; (iii) cytoplasm-nuclear traffic by exposing/masking nuclear localization signals, or by releasing the transcription factor from partners or from membrane anchors; and, (iv) DNA binding and nuclear transactivation by modulating transcription factor affinity towards DNA, co-activators or repressors, and by targeting specific regions of chromatin to activate individual genes. We also discuss how H2O2 biological specificity results from diverse thiol protein sensors, with different reactivity of their sulfhydryl groups towards H2O2, being activated by different concentrations and times of exposure to H2O2. The specific regulation of local H2O2 concentrations is also crucial and results from H2O2 localized production and removal controlled by signals. Finally, we formulate equations to extract from typical experiments quantitative data concerning H2O2 reactivity with sensor molecules. Rate constants of 140 M-1s−1 and ≥ 1.3 × 103 M-1s−1 were estimated, respectively, for the reaction of H2O2 with KEAP1 and with an unknown target that mediates NRF2 protein synthesis. In conclusion, the multitude of H2O2 targets and mechanisms provides an opportunity for highly specific effects on gene regulation that depend on the cell type and on signals received from the cellular microenvironment.

Insights into the role of almond CBF transcription factors in the environmental control of cold acclimation and dormancy break

Dissertation presented to obtain the Ph.D degree in Biology

Regulation of the rice gene OsRMC under salt stress: identification and functional characterization of novel transcription factors

Dissertation presented to obtain the Ph.D degree in Biology