Endoplasmic Reticulum-Associated Degradation (ERAD)

The newly synthesized proteins are kept in the endoplasmic reticulum (ER) until their maturation is completed. The accurate protein folding is vital for homeostasis, but this process is error-prone since it is chemically complicated. Aberrant folding may result in aggregates having a toxic gain of function or may lead to a loss of protein function; therefore, protein misfolding can lead to several pathologies. The ER protein quality control mechanism monitors the fidelity of protein folding. Those proteins that fail to fold or assemble properly are subjected to degradation via a process known as ER-associated degradation (ERAD). Besides clearing proteins having folding problems, ERAD is also known to regulate the levels of some physiological proteins including 3-hydroxy-3-methylglutaryl-coenzymeA reductase (HMGR) catalyzing the rate-limiting step of cholesterol biosynthesis. ERAD is a complex, multistep process starting with the recognition and targeting of substrates, followed by ubiquitination, retrotranslocation and proteasomal degradation. A large number of ERAD factors functioning in different molecular machineries increases the complexity of mammalian ERAD. ERAD is fundamental for human health and there is increasing evidence linking ERAD with various diseases. Here, the different modules/machineries of the ERAD process together with its tight regulation will be discussed

DNA Hasarı ve onarımında NKX3.1'in rolü

Bu tez çalışmasında, prostat kanseri hücre hatlarında topoizomeraz inhibitörleri ve oksidatif stres ajanları kullanılarak, topoizomeraz-I aktivitesini arttıran bir gen olarak NKX3.1'in DNA hasarı yanıtı ve onarımındaki rolü araştırılmıştır. CPT-11 uygulaması ile yaratılan DNA hasarına yanıt olarak artan H2AX(S139) odak sayılarının ve toplam fosforilasyon seviyelerinin, PC-3 ve DU145 hücrelerinde kazandırılmış NKX3.1 ekspresyonu ve LNCaP hücrelerinde androjen uygulaması ile birlikte azaldığı bulgulanmıştır. NKX3.1 varlığında siklin D1 ekspresyon seviyesinin madde uygulamasından bağımsız olarak yüksek seviyede olduğu belirlenmiştir. Ayrıca, immünopresipitasyon yöntemi kullanılarak NKX3.1'in RAD50, DNA-PKcs ve p-ATM(S1981) gibi DNA hasarı onarım proteinleri ile etkileştiği saptanmıştır. Ek olarak, NKX3.1 E63-Cter mutant proteininin ne ÞH2AX(S139) ne de p-ATM(S1981) ile bir etkileşim göstermediği ve bu nedenle NKX3.1 homeodomain bölgesi yokluğuna bağlı olarak kaybolan NKX3.1 işlev kaybı durumunda ÞH2AX(S139) odak oluşumlarının geri döndüğü gözlenmiştir. Son olarak PC-3 ve LNCaP hücrelerinde NKX3.1'in topoizomeraz inhibitörlerine karşı oluşturulan yanıtın yanı sıra oksidatif stres aracılıklı DNA hasarına karşı oluşturulan yanıtta da oldukça önemli olduğu ve ektopik NKX3.1 ekspresyonu ya da androjenlerin (uyarılmış NKX3.1 ekspresyonu) varlığında DNA hasarına alınan yanıtların değiştiği saptanmıştır. Tüm bu verilerden yola çıkarak, androjen veya NKX3.1'in varlığında hücrede hasar algısının arttığı ve onarımın daha hızlı bir şekilde gerçekleştiği, NKX3.1 yokluğunda ise onarım komplekslerinin yeterince aktif olamadığı ve hasarın biriktiği sonucuna varılmıştır

Disodium pentaborate decahydrate (DPD) induced apoptosis by decreasing hTERT enzyme activity and disrupting F-actin organization of prostate cancer cells

WOS: 000332026300087PubMed ID: 24122279Animal and cell culture studies have showed that boron and its derivatives may be promising anticancer agents in prostate cancer treatment. Thus, DU145 cells were treated with disodium pentaborate decahydrate (DPD) for 24, 48, and 72 h in order to investigate the inhibitor effect and mechanisms of DPD. Then, cell proliferation, telomerase enzyme activity, actin polymerization, and apoptosis were detected by WST-1 assay, qRT-PCR, immunofluorescence labeling, and flow cytometry, respectively. We found that DPD inhibited the growth of human prostate cancer cell line DU145 at the concentration of 3.5 mM for 24 h. Our results demonstrated that 7 mM of DPD treatment prevented the telomerase enzyme activity at the rate of 38 %. Furthermore, DPD has an apoptotic effect on DU145 cells which were examined by labeling DNA breaks. With 7 mM of DPD treatment, 8, 14, and 41 % of apoptotic cells were detected for 24, 48, and 72 h, respectively. Additionally, immunofluorescence labeling showed that the normal organization of actin filaments was disrupted in DPD-exposed cells, which is accompanied by the alteration of cell shape and by apoptosis in targeted cells. Taken together, the results indicate that DPD may exert its cytotoxicity at least partly by interfering with the dynamic properties of actin polymerization and decreasing the telomerase activity. Eventually, for the first time, the results of this study showed that DPD suppressed the activity of telomerase in DU145 cells, and therefore, we suggested that DPD could be an important agent for its therapeutic potential in the treatment of prostate cancer.BAP projects by the Celal Bayar UniversityCelal Bayar University [2009-92, 2010-91]We would like to thank the National Boron Research Institute, BOREN, for providing DPD. This research was supported with grants (2009-92 and 2010-91) from BAP projects by the Celal Bayar University

Exploring of tumor-associated carbonic anhydrase isoenzyme IX and XII inhibitory effects and cytotoxicities of the novel N-aryl-1-(4-sulfamoylphenyl)-5-(thiophen-2-yl)-1H-pyrazole-3-carboxamides

A series of novel N-aryl-1-(4-sulfamoylphenyl)-5-(thiophen-2-yl)-1H-pyrazole-3-carboxamides was synthesized and examined as inhibitors of cytosolic (human) hCA I and hCA II, and cancer-related transmembrane hCA IX and hCA XII isoenzymes. AC2 was the most selective inhibitor towards cancer-related hCA IX while AC8 and AC9 selectively inhibited hCA XII over off-target isoenzymes. Anticancer effects of the compounds were evaluated towards human oral squamous cell carcinoma (OSCC) cell lines, human mesenchymal normal oral cells, breast (MCF7), prostate (PC3), non-small cell lung carcinoma cells (A549), and non-tumoral fetal lung fibroblast cells (MRC5). Compounds moderately showed cytotoxicity towards cancer cell lines. Among others, AC6 showed cell specific cytotoxic activity and induced apoptosis in a dose-dependent manner without a significant change in the cell cycle distribution of MCF7. These results suggest that pyrazole-3-carboxamides need further molecular modification to increase their anticancer drug candidate potency.Scientific and Technological Research Council of Turkey (TUBITAK) [219S076]We gratefully acknowledge financial support from The Scientific and Technological Research Council of Turkey (TUBITAK, Project no: 219S076)

Prostat kanseri hücrelerinde MAP kinaz gen ekspresyonlarını yukarı doğru regüle eden disodyum pentaborat dekahidrat

Ege Üniversitesi, Tıp FakültesiAvrasya Üniversitesi, Fen - Edebiyat FakültesiPamukkale <üniversitesi, Tıp FakültesiCelal Bayar Üniversitesi, Tıp FakültesiObjectives: This study aims to investigate the expressions of MAP2K3, MAP3K7, and MAPK8 genes after disodium pentaborate decahydrate (DPD) treatment in DU-145 cells. Materials and methods: Effect of DPD treatment on expressions of MAP2K3, MAP3K7, and MAPK8 genes were determined by qRT-PCR. Results: We determined that disodium pentaborate decahydrate treatment increased the expressions of MAP2K3, MAP3K7, and MAPK8 genes in terms of mRNA levels. Conclusion: The reason of increased apoptosis might be associated with high expression levels of MAP2K3, MAP3K7, and MAPK8 genes after DPD treatment. Our novel findings suggest that DPD may be an important agent in the treatment of prostate cancer by inducing apoptosis against mitogenic and environmental stress.Amaç: Bu çalışmada DU-145 hücrelerinde disodyum pentaborat dekahidrat (DPD) uygulamasının ardından MAP2K3, MAP3K7 ve MAPK8 genlerinin ekspresyonları araştırıldı. Gereç ve yöntemler: Disodyum pentaborat dekahidrat uygulamasının MAP2K3, MAP3K7 ve MAPK8 genlerinin ekspresyonlarına olan etkisi qRT-PCR ile belirlendi. Bulgular: Disodyum pentaborat dekahidrat uygulamasının MAP2K3, MAP3K7 ve MAPK8 genlerinin mRNA seviyelerini artırdığı belirlendi. Sonuç: Apoptoz artışının, MAP2K3, MAP3K7 ve MAPK8 genlerinin DPD uygulaması sonrasında yükselen ekspresyon seviyelerine bağlı olarak gerçekleştiği düşünülmektedir. Güncel bulgularımız DPD’nin prostat kanseri tedavisinde mitojenik ve çevresel strese karşı apoptoza neden olan önemli bir etken olabileceğini öngörmektedir