To maintain intracellular redox homeostasis, genes encoding many endogenous antioxidants and phase II detoxification enzymes are transcriptionally upregulated upon deleterious oxidative stress through the cis- antioxidant responsive elements (AREs) in their promoter regions. Nrf2 has emerged as the pivatol transcription factor responsible for ARE-dependent transcription, and has been shown to play critical roles in hepatotoxicity, chemical carcinogenesis, pulmonary inflammatory diseases, neurodegenerative diseases and aging. Therefore, understanding the molecular mechanism of the Nrf2-dependent cytoprotective system is important for development of drugs for therapeutic intervention.Nrf2 is targeted by Keap1 for ubiquitin-mediated degradation under basal conditions. Upon oxidative stress, distinct cysteine residues of Keap1 are alkylated, leading to inhibition of Keap1 and activation of Nrf2. However, it was not clear how Nrf2 is re-entered into the repression status when redox homeostasis is re-achieved. In this dissertation, we establish that the post-induction repression of Nrf2 is controlled by the nuclear export function of Keap1 in alliance with the cytoplasmic ubiquitination/ degradation machinery. We show that a nuclear export sequence (NES) in Keap1 is required for termination of Nrf2 signaling; ubiquitination of Nrf2 is carried out in the cytosol; Keap1 nuclear translocation is independent of Nrf2; and the Nrf2-Keap1 complex does not bind the ARE. Collectively, these results suggest that Keap1 translocates into the nucleus to dissociate Nrf2 from the ARE and mediates nuclear export of Nrf2 followed by ubiquitination and degradation of Nrf2 in the cytoplasm.In addition to Keap1-mediated negative regulation, we identified a novel positive regulatory mechanism of Nrf2 mediated by transcription co-activator p300/CBP. We show that p300/CBP directly binds and acetylates Nrf2 in response to oxidative stress. We have identified multiple acetylated lysine residues within the Nrf2 Neh1 DNA-binding domain. Combined lysine-to-arginine mutations on the acetylation sites, with no effects on Nrf2 protein stability, compromised the DNA-binding activity of Nrf2 in a promoter-specific manner both in vitro and in vivo. These findings demonstrated that acetylation of Nrf2 by p300/CBP augments promoter-specific DNA binding of Nrf2 and established acetylation as a novel regulatory mechanism that functions in concert with Keap1-mediated ubiquitination in modulating the Nrf2-dependent antioxidant response.

Objective: P300/CBP Associated Factor (PCAF) is a histone acetyltransferase, and has been reported to interact with nuclear factor κ-light-chain-enhancer of activated B cells (NF-κB) and to stimulate cyclooxygenase-2 (COX-2) transcriptional activation. The aim of this study was to determine the role of PCAF in chronic pain modulation. / Method: In an in vitro experiment, PCAF small-interfering RNA (siRNA) was used to knock down PCAF. Interleukin-1 β (IL1β) was applied as COX-2 inducer in SK-N-SH neuroblastoma cells. Luciferase assay and chromatin immunoprecipitation (ChIP) were performed regarding COX-2 promoter region. / In an in vivo experiment, PCAF was examined for its distribution in dorsal horn of Spraque-Dawley rats. COX-2 level in the spinal cord was determined after inhibition of PCAF in rats with Complete Freund's Adjuvant (CFA)-induced pain. ChIP was also performed. / Finally, we tested whether genetic variations in the PCAF gene affected the risk of chronic pain in a gene association study of 267 surgical patients. The associations of pain with genotypes (58 single nucleotide polymorphisms, SNPs)/haplotypes were analyzed by χ² and Fisher exact tests. / Results: Knock-down of PCAF reduced COX-2 level and NF-κB activity. PCAF and acetylated histone 3 lysine 14 (H3K14) were enriched at COX-2 promoter when IL1β was applied. / PCAF was expressed in neurons at superficial layers of rat dorsal horn. In the in vivo experiment, COX-2 was reduced with the inhibition of PCAF in CFA rats. PCAF was enhanced at COX-2 promoter when CFA was injected. Anacardic acid and PCAF siRNA significantly alleviated thermal nociception and mechanical allodynia. / In the gene association study, 6 SNPs and 5 haplotypes were significantly associated with higher risk of severe chronic postsurgical pain. Multivariable analysis showed that patients with a SNP rs6763504 had a higher risk of developing severe chronic postsurgical pain (p = 0.001). / Conclusion: PCAF regulated COX-2 transcription and reduction or inhibition in PCAF resulted in a decrease in COX-2 and less chronic inflammatory pain. Genetic variations in the PCAF gene increased risk of severe chronic post-surgical pain in patients. / 實驗目的：P300/CBP相關蛋白（PCAF）是一種組蛋白乙酰化轉移酶。這種蛋白已被報道可以和核因子活化B細胞κ輕鏈增強子(NF-κB)相互作用，以及增進環氧合酶-2（COX-2）的轉錄激活。本實驗的目的在於研究PCAF在疼痛調節中的作用。 / 實驗方法：在細胞實驗中，我們使用了小干擾RNA（siRNA）來降低PCAF的含量。同時我們使用了白細胞介素-1β（IL1β）來作為SK-N-SH神經母細胞瘤細胞中COX-2的誘導劑。我們使用了熒光素酶試劑和染色質免疫沉澱來研究COX-2啟動子區域。 / 在體內實驗中，我們檢測了PCAF在大鼠脊椎背角部位的分佈。在弗氏完全佐劑（CFA）致痛的大鼠模型中，我們在PCAF抑制的情況下檢測脊髓中COX-2的水平。我們還進行了染色質免疫沉澱。 / 最後，在招募了267位手術患者的基因關聯試驗中，我們對PCAF基因中的基因變異對慢性疼痛風險的影響進行了分析。我們運用卡方檢驗和費雪精確性檢驗對疼痛與基因型（58個單核苷酸多態性）和單倍型的關係進行分析。 / 實驗結果：減少的PCAF降低了COX-2的水平以及NF-κB的活性。當添加了IL1β時PCAF和乙酰化第三組蛋白14號賴胺酸（H3K14）在COX-2啟動子處富集。 / PCAF在大鼠脊椎背角部位的淺表層（第一層和第二層）的神經細胞里表達。在動物實驗中，注射了CFA的大鼠顯示COX-2會隨著PCAF的抑制而下降。大鼠注射了CFA后PCAF在COX-2啟動子處有所增加。漆樹酸和PCAF siRNA顯著地減輕了熱痛和機械痛提高了機械痛閾值。 / 在該基因關聯試驗中，我們鑒定出六個單核苷酸多態性和五種單倍型與較高風險的嚴重術後慢性痛有顯著的相關性。多元回歸分析表明在PCAF基因上具有rs6763504遺傳變異的病人在術後發展嚴重慢性痛的幾率會較高(p = 0.001)。 / 結論：PCAF調節COX-2的轉錄，而且PCAF的減少或者抑制導致了COX-2的降低同時慢性炎症痛的減少。PCAF基因上的遺傳變異提高了術後病人嚴重慢性痛的風險。 / Meng, Zhaoyu. / Thesis (Ph.D.)--Chinese University of Hong Kong, 2014. / Includes bibliographical references (leaves 117-134). / Abstracts also in Chinese. / Title from PDF title page (viewed on 30, December, 2016). / Detailed summary in vernacular field only. / Detailed summary in vernacular field only. / Detailed summary in vernacular field only. / Detailed summary in vernacular field only. / Detailed summary in vernacular field only. / Detailed summary in vernacular field only. / Detailed summary in vernacular field only. / Detailed summary in vernacular field only.

Thesis (Ph. D.)--Case Western Reserve University, 2006. / [School of Medicine] Department of Pharmacology. Includes bibliographical references. Available online via OhioLINK's ETD Center.

Posttranslational modifications are of great importance in the mediation of transcriptional effects, necessary for signalling in cancer. A characteristic example of such modifications is acetylation of the p53 tumour suppressor, a transcription factor involved in several crucial cellular functions including cell-cycle arrest and apoptosis. p53 is stabilised under hypoxic and DNA damaging-conditions. However, only in the latter scenario is p53 fully capable of inducing the expression of its proapoptotic targets through acetylation. The hypoxia inducible factor 1 (HIF-1) transcription factor is stabilised at low oxygen levels to mediate a cellular adaptive response under these conditions, promoting cell survival. As these two opposing transcription factors share a common transcriptional regulator, p300/CBP, this study focused on deciphering the p300/CBP complex components under differential stress to determine its composition required for cellular responses elicited in response to DNA damage or hypoxia, in an effort to investigate a possible link between differential posttranslational modifications and the resulting cell fate. Hence, the aim of this study was to investigate the roles of p300/CBP components in dictating transcriptional regulation of both HIF-1 and p53 in hypoxic conditions. To carry out this study, the proapoptotic BID gene was the system used, as its promoter contains a p53 response element and a HIF-1 response element (HRE). The p300/CBP associated factors PCAF and Strap were appointed as potent candidates for posttranslational modifications under differential conditions, as they are stress-responsive cofactors. Under DNA damage, PCAF acetylates p53 at K320 and Strap augments p300 binding to p53, both of which amplify the p53 response. Evidence from this study demonstrates that under hypoxia-mimicking conditions PCAF-mediated p53 acetylation at K320 is reduced to a greater extent compared to p300/CBP acetylation at K382. The limited amounts of acetylated p53 at K320 are preferentially recruited to the promoter of the cell cycle arrest p21WAF-1/CIP-1 gene that appears to be unaffected by hypoxia, but fail to be recruited to the BID promoter, rendering p53 incapable of upregulating proapoptotic BID in hypoxic conditions. In addition, under the same conditions, PCAF was found to acetylate, and direct HIF-1 to a particular subset of its targets, leading to alterations in the net physiological effect. Moreover, the intrinsic acetyl transferase activity of PCAF was shown to increase the stability of HIF-1. An additional role was attributed to PCAF in relation to apoptosis, albeit from another angle. BID protein translocation to the cytoplasm in hypoxic conditions was facilitated by ectopically expressed PCAF.Strap was found to be preferentially recruited to the HRE of the BID promoter in hypoxic conditions, and to exert a transrepression effect that appeared to be p53-dependent. Strap also interacted with specific PCAF isoforms depending on the type of cellular stress. Contrary to PCAF, ectopically expressed Strap did not have any effect on BID subcellular distribution. This study has provided additional insight in the mechanisms by which cofactors are involved in cell fate, either by affecting activity and stability of HIF-1 and p53, or having a direct effect on Bcl-2 member subcellular distribution.

For 30 years, the tumor suppressor p53 has been a subject of intense research in nearly every discipline of scientific inquiry. While numerous surprising roles for p53 in health and disease are uncovered each year, the central role of its activation in preventing neoplastic transformation has been and will remain at the forefront of p53 research as investigators work to address an unexpectedly complex question—precisely how does p53 integrate upstream stress signals to coordinate activation of its target genes in response to stress? One manner in which to address this question is at the level of transcription initiation—after upstream signals converge on p53 and produce a number of pools of post-transcriptionally modified p53, how exactly are specific target promoters activated in such a sensitive, context-specific manner? The work presented herein aims to address the role of histone acetylation at the p21 promoter—a critical mediator of G1/S arrest—by the P/CAF acetyltransferase in response to a variety of p53-activating stresses. We show that depletion of P/CAF strongly inhibits p21 expression in response to a variety of stresses, despite normal stabilization of p53 and recruitment to target promoters. This defect in p21 expression correlates closely with abrogation of stress-induced cell-cycle arrest. Strikingly, a p53 allele lacking putative P/CAF acetylation sites was still able to direct p21 expression, which was still dependent upon P/CAF. We show further that histone acetylation at H3K14 at the p21 promoter following stress is dependent upon P/CAF. Rescue of p21 expression with wild-type P/CAF or a ∆HAT point mutant indicates that P/CAF requires an intact HAT domain, suggesting that histone acetylation at H3K14 is catalyzed by P/CAF HAT activity, not the molecular bridging of a heterologous HAT by P/CAF. Furthermore, RNA polymerase II (RNAP II) was present at the p21 proximal promoter under all basal and stress conditions, but elongation of RNAP II after stress required the presence of P/CAF. These data indicate that H3K14 acetylation by P/CAF closely correlates with the activation status of the p21 promoter, and may be necessary for activation of a larger subset of p53-responsive promoters. In addition to its critical role in p21 expression, we noted that p53 stabilization and cell-cycle arrest in response to p14ARF, but not other p53-stabilizing stresses, were also dependent on P/CAF. Cell-cycle arrest induced by p16INK4A was intact after P/CAF ablation, indicating a role for P/CAF in cell-cycle arrest specific to p14ARF-p53 signaling. Basal MDM2 levels were unaffected by P/CAF knockdown, as were p53- MDM2 and ARF-MDM2 complexes. A preliminary analysis of MDM2 localization was inconclusive, due to vastly different quantities of MDM2 in different conditions making analysis of subcellular localization difficult; however, the role of P/CAF in the relocalization of MDM2 to the nucleolus by p14ARF could potentially explain the defect in p53 stabilization, and should be explored further. These observations, underscored by recent reports that P/CAF undergoes loss of heterozygosity in several tumor types, suggest that P/CAF plays a critical role in p53-mediated cell-cycle arrest through multiple, independent mechanisms. Further study should clarify whether P/CAF is lost in tumors maintaining wild-type p53, and whether its reintroduction into these tumors confers any potential therapeutic benefit.

INTRODUÇÃO: A Síndrome de Rubinstein-Taybi (RTS) é uma doença genética rara, caracterizada por dismorfismos craniofaciais típicos, polegares e háluces alargados, deficiência mental e baixa estatura. A incidência estimada é de 1: 125 000 a 1: 330000 nativivos. A SRT geralmente ocorre esporadicamente, mas pode ser herdada com um padrão de herança autossômico dominante. O diagnóstico da SRT é essencialmente clínico. OBJETIVOS: Realizar o estudo genético-clínico e citogenético em 30 pacientes brasileiros com SRT, e descrever de forma sistematizada a freqüência de dismorfismos faciais e malformações múltiplas encontradas. MÉTODOS: Neste estudo observacional retrospectivo e prospectivo, os pacientes foram seguidos no período de agosto de 2005 a junho de 2009. O cariótipo com bandeamento G foi realizado em todos os pacientes. RESULTADOS: A maioria dos pacientes avaliados foi do sexo feminino (60%). As seguintes características foram observadas em todos os pacientes da nossa casuística: atraso de desenvolvimento neuropsicomotor, ponta nasal voltada para baixo, columela proeminente, sorriso característico, dificuldades alimentares na infância, persistência dos coxins fetais, falanges distais dos polegares alargadas e pés planos. A baixa estatura e a microcefalia foi observada em 80% e 76% dos casos, respectivamente. As principais características craniofaciais observadas foram: fronte proeminente (86%), ponte nasal larga (60%), hipertelorismo (70%), sobrancelhas espessas e arqueadas (96%) cílios longos em 93%, prega epicântica (76%), fissura palpebral infra vertidas (76%), abertura bucal estreita (93%), retrognatismo (76 %), sorriso característico em 100%, palato alto e estreito (93%), anomalias dentárias (83%). Outras anomalias identificadas foram: estrabismo, erros de refração, obstrução do canal lacrimal, háluces e polegares alargados, angulação de polegares, anomalias do pavilhão auricular (rotação/posição/tamanho/forma), angulação do hálux, clinodactilia, sobreposição dos pododáctilos, falanges distais alargadas de outros dedos, marcha rígida, hipotonia, sopro cardíaco, cardiopatia congênita, criptoquidia, hemangioma plano e hipertricose. Uma paciente apresentou translocação recíproca de novo 46, XX, t (2; 16)(q36.3; p13.3). CONCLUSÕES: A raridade da SRT e o amplo espectro das manifestações clínicas pode atrasar o diagnóstico clínico. A média da idade ao diagnóstico dos nossos pacientes com SRT foi de três anos e oito meses. Todas as crianças devem receber avaliação por geneticista pediátrico, cardiologista, oftalmologista, neuropediatra, e odontopediatra / INTRODUCTION: Rubinstein-Taybi syndrome (RTS) is a rare genetic disorder characterized by distinctive craniofacial dysmorphisms, broad thumbs and toes and mental and statural deficiency. The prevalence of RTS has been estimated to be 1 in 125000 to 1: 330000 live births. RTS usually occurs sporadically although it can be inherited as an autosomal dominant disorder. The diagnosis of RTS is primarily based on clinical features. OBJECTIVES: We performed a clinical and cytogenetic assay in a group of 30 Brazilian RTS patients. We also decribed the frequencies of facial dysmorphisms and multiple malformations. METHODS: In this observational retrospective and prospective study, the patients were followed from August 2005 to June 2009. Chromosomal analysis was performed by G-banding karyotype. RESULTS: Most of the patients were female (60%).The following abnormalities were present in all of the patients: delayed psychomotor development, beaked nose, proeminent collumel, typical facies, broad thumbs and toes, flat feet, joint laxity, feeding problems during the childhood, and finger pads. Short stature was present in 80%, and microcephalia in 76% of the cases, respectively. Main craniofacial characteristics are frontal bossing (86%), wide nasal bridge (60%), ocular hyperthelorism (70%), high arched eyebrows (96%), long eyelashes (93%), epicathal folds (76%), downslanting palpebral fissures (76%), small opening of the mouth (93%), retrognathism (76%), grimacing smile (100%), high arched palate (93%), and dental anomalies (83%). Other findings were: strabism, refractive error, lacrimal obstruction, wide thumb and halux, angulated thumbs, external ears anomalies (rotation, implantation and morfology), angulated halux, clinodactyly, crowded toes, broad distal falanges of other fingers, stiff gait, hipotonia, cardiac murmur, congenital heart defect, undescendent testis, hypertrichosis, and hemangioma. One female patient has found to have a reciprocal de novo translocation t(2;16)(q36.3;p13.3) on G-banding karyotype CONCLUSIONS: The rarity of RTS and the wide spectrum of clinical manifestations, may delay the clinical diagnosis of RTS. The average age at the diagnosis of our patients was 3 years and 8 months. All children of RTS should receive an evaluation by a pediatric geneticist, cardiologist, ophthalmologist, pediatric neurologist, and pediatric dentist

INTRODUÇÃO: A Síndrome de Rubinstein-Taybi (RTS) é uma doença genética rara, caracterizada por dismorfismos craniofaciais típicos, polegares e háluces alargados, deficiência mental e baixa estatura. A incidência estimada é de 1: 125 000 a 1: 330000 nativivos. A SRT geralmente ocorre esporadicamente, mas pode ser herdada com um padrão de herança autossômico dominante. O diagnóstico da SRT é essencialmente clínico. OBJETIVOS: Realizar o estudo genético-clínico e citogenético em 30 pacientes brasileiros com SRT, e descrever de forma sistematizada a freqüência de dismorfismos faciais e malformações múltiplas encontradas. MÉTODOS: Neste estudo observacional retrospectivo e prospectivo, os pacientes foram seguidos no período de agosto de 2005 a junho de 2009. O cariótipo com bandeamento G foi realizado em todos os pacientes. RESULTADOS: A maioria dos pacientes avaliados foi do sexo feminino (60%). As seguintes características foram observadas em todos os pacientes da nossa casuística: atraso de desenvolvimento neuropsicomotor, ponta nasal voltada para baixo, columela proeminente, sorriso característico, dificuldades alimentares na infância, persistência dos coxins fetais, falanges distais dos polegares alargadas e pés planos. A baixa estatura e a microcefalia foi observada em 80% e 76% dos casos, respectivamente. As principais características craniofaciais observadas foram: fronte proeminente (86%), ponte nasal larga (60%), hipertelorismo (70%), sobrancelhas espessas e arqueadas (96%) cílios longos em 93%, prega epicântica (76%), fissura palpebral infra vertidas (76%), abertura bucal estreita (93%), retrognatismo (76 %), sorriso característico em 100%, palato alto e estreito (93%), anomalias dentárias (83%). Outras anomalias identificadas foram: estrabismo, erros de refração, obstrução do canal lacrimal, háluces e polegares alargados, angulação de polegares, anomalias do pavilhão auricular (rotação/posição/tamanho/forma), angulação do hálux, clinodactilia, sobreposição dos pododáctilos, falanges distais alargadas de outros dedos, marcha rígida, hipotonia, sopro cardíaco, cardiopatia congênita, criptoquidia, hemangioma plano e hipertricose. Uma paciente apresentou translocação recíproca de novo 46, XX, t (2; 16)(q36.3; p13.3). CONCLUSÕES: A raridade da SRT e o amplo espectro das manifestações clínicas pode atrasar o diagnóstico clínico. A média da idade ao diagnóstico dos nossos pacientes com SRT foi de três anos e oito meses. Todas as crianças devem receber avaliação por geneticista pediátrico, cardiologista, oftalmologista, neuropediatra, e odontopediatra / INTRODUCTION: Rubinstein-Taybi syndrome (RTS) is a rare genetic disorder characterized by distinctive craniofacial dysmorphisms, broad thumbs and toes and mental and statural deficiency. The prevalence of RTS has been estimated to be 1 in 125000 to 1: 330000 live births. RTS usually occurs sporadically although it can be inherited as an autosomal dominant disorder. The diagnosis of RTS is primarily based on clinical features. OBJECTIVES: We performed a clinical and cytogenetic assay in a group of 30 Brazilian RTS patients. We also decribed the frequencies of facial dysmorphisms and multiple malformations. METHODS: In this observational retrospective and prospective study, the patients were followed from August 2005 to June 2009. Chromosomal analysis was performed by G-banding karyotype. RESULTS: Most of the patients were female (60%).The following abnormalities were present in all of the patients: delayed psychomotor development, beaked nose, proeminent collumel, typical facies, broad thumbs and toes, flat feet, joint laxity, feeding problems during the childhood, and finger pads. Short stature was present in 80%, and microcephalia in 76% of the cases, respectively. Main craniofacial characteristics are frontal bossing (86%), wide nasal bridge (60%), ocular hyperthelorism (70%), high arched eyebrows (96%), long eyelashes (93%), epicathal folds (76%), downslanting palpebral fissures (76%), small opening of the mouth (93%), retrognathism (76%), grimacing smile (100%), high arched palate (93%), and dental anomalies (83%). Other findings were: strabism, refractive error, lacrimal obstruction, wide thumb and halux, angulated thumbs, external ears anomalies (rotation, implantation and morfology), angulated halux, clinodactyly, crowded toes, broad distal falanges of other fingers, stiff gait, hipotonia, cardiac murmur, congenital heart defect, undescendent testis, hypertrichosis, and hemangioma. One female patient has found to have a reciprocal de novo translocation t(2;16)(q36.3;p13.3) on G-banding karyotype CONCLUSIONS: The rarity of RTS and the wide spectrum of clinical manifestations, may delay the clinical diagnosis of RTS. The average age at the diagnosis of our patients was 3 years and 8 months. All children of RTS should receive an evaluation by a pediatric geneticist, cardiologist, ophthalmologist, pediatric neurologist, and pediatric dentist

p53, known as the “guardian of the genome”, is the most well-characterized tumor suppressor gene. The central role of p53 is to prevent genome instability. p53 is the central node in an incredibly elaborate genome defense network for receiving various input stress signals and controlling diverse cellular responses. The final output of this network is determined not only by the p53 protein itself, but also by other p53 cooperating proteins. p300 and CBP (CREB-Binding Protein) act as multifunctional regulators of p53 via acetylase and ubiquitin ligase activities. Prior work in vitro has shown that the N-terminal 595 aa of p300 encode both generic ubiquitin ligase (E3) and p53-directed E4 functions. Analysis of p300 or CBP-deficient cells revealed that both coactivators were required for endogenous p53 polyubiquitination and the normally rapid turnover of p53 in unstressed cells. Unexpectedly, p300/CBP ubiquitin ligase activities were absent in nuclear extracts and exclusively cytoplasmic. In the nucleus, CBP and p300 exhibited differential regulation of p53 gene target expression, C-terminal acetylation, and biologic response after DNA damage. p300 activated, and CBP repressed, PUMA expression, correlating with activating acetylation of p53 C-terminal lysines by p300, and a repressive acetylation of p53 lysine-320 induced by CBP. Consistent with their gene expression effects, CBP deficiency augmented, and p300 deficiency blocked, apoptosis after doxorubicin treatment. Subcellular compartmentalization of p300/CBP’s ubiquitination and transcription activities reconciles seemingly opposed functions—cytoplasmic p300/CBP E4 activities ubiquitinate and destabilize p53, while nuclear p300/CBP direct p53 acetylation, target gene activation, and biological outcome after genotoxic stress. p53 is a prominent tumor suppressor gene and it is mutated in more than 50% of human tumors. Reactivation of endogenous p53 is one therapeutic avenue to stop cancer cell growth. In this thesis, we have identified S5as a critical regulator of p53 degradation and activity. S5a is a non-ATPase subunit in the 19S regulatory particle of the 26S proteasome. Our preliminary data indicates that S5a is required for p53 instability and is a negative regulator of p53 tranactivation. As a negative regulator of p53, S5a may therefore also represent a new target for cancer drug development against tumors that specifically maintain wild type p53.

No / Resveratrol is a polyphenolic phytoestrogen that has been shown to exhibit potent anti-oxidant, anti-inflammatory, and anti-catabolic properties. Increased osteoclastic and decreased osteoblastic activities result in bone resorption and loss of bone mass. These changes have been implicated in pathological processes in rheumatoid arthritis and osteoporosis. Receptor activator of NF-kappaB ligand (RANKL), a member of the TNF superfamily, is a major mediator of bone loss. In this study, we investigated the effects of resveratrol on RANKL during bone morphogenesis in high density bone cultures in vitro. Untreated bone-derived cell cultures produced well organized bone-like structures with a bone-specific matrix. Treatment with RANKL induced formation of tartrate-resistant acid phosphatase-positive multinucleated cells that exhibited morphological features of osteoclasts. RANKL induced NF-kappaB activation, whereas pretreatment with resveratrol completely inhibited this activation and suppressed the activation of IkappaBalpha kinase and IkappaBalpha phosphorylation and degradation. RANKL up-regulated p300 (a histone acetyltransferase) expression, which, in turn, promoted acetylation of NF-kappaB. Resveratrol inhibited RANKL-induced acetylation and nuclear translocation of NF-kappaB in a time- and concentration-dependent manner. In addition, activation of Sirt-1 (a histone deacetylase) by resveratrol induced Sirt-1-p300 association in bone-derived and preosteoblastic cells, leading to deacetylation of RANKL-induced NF-kappaB, inhibition of NF-kappaB transcriptional activation, and osteoclastogenesis. Co-treatment with resveratrol activated the bone transcription factors Cbfa-1 and Sirt-1 and induced the formation of Sirt-1-Cbfa-1 complexes. Overall, these results demonstrate that resveratrol-activated Sirt-1 plays pivotal roles in regulating the balance between the osteoclastic versus osteoblastic activity result in bone formation in vitro thereby highlighting its therapeutic potential for treating osteoporosis and rheumatoid arthritis-related bone loss.

No / Vitiligo is characterized by a patchy loss of inherited skin color affecting approximately 0.5% of individuals of all races. Despite the absence of the protecting pigment and the overwhelming evidence for hydrogen peroxide (H(2)O(2))-induced oxidative stress in the entire epidermis of these patients, there is neither increased photodamage/skin aging nor a higher incidence for sun-induced nonmelanoma skin cancer. Here we demonstrate for the first time increased DNA damage via 8-oxoguanine in the skin and plasma in association with epidermal up-regulated phosphorylated/acetylated p53 and high levels of the p53 antagonist p76(MDM2). Short-patch base-excision repair via hOgg1, APE1, and polymerasebeta DNA repair is up-regulated. Overexpression of Bcl-2 and low caspase 3 and cytochrome c levels argue against increased apoptosis in this disease. Moreover, we show the presence of high epidermal peroxynitrite (ONOO(-)) levels via nitrotyrosine together with high nitrated p53 levels. We demonstrate by EMSA that nitration of p53 by ONOO(-) (300 x 10(-6) M) abrogates DNA binding, while H(2)O(2)-oxidized p53 (10(-3) M) enhances DNA binding capacity and prevents ONOO(-)-induced abrogation of DNA binding. Taken together, we add a novel reactive oxygen species to the list of oxidative stress inducers in vitiligo. Moreover, we propose up-regulated wild-type p53 together with p76(MDM2) as major players in the control of DNA damage/repair and prevention of photodamage and nonmelanoma skin cancer in vitiligo.