Browsing by Author "Adacan, Kaan"

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Calreticulin is a fine tuning molecule in epibrassinolide-induced apoptosis through activating endoplasmic reticulum stress in colon cancer cells
(Wiley, 111 River St, Hoboken 07030-5774, NJ USA, 2017-06) Çoker Gürkan, Ajda; Adacan, Kaan; Özbey, Utku; Somuncu, Berna; Baran, Didem; Ünsal Palavan, Zeynep Narçın; ARISAN, ELİF DAMLA; YERLİKAYA, PINAR OBAKAN; 156421; 113920; 125860; 6125
Epibrassinolide (EBR), a member of brassinostreoids plant hormones with cell proliferation promoting role in plants, is a natural polyhydroxysteroid with structural similarity to steroid hormones of vertebrates. EBR has antiproliferative and apoptosis-inducing effect in various cancer cells. Although EBR has been shown to affect survival and mitochondria-mediated apoptosis pathways in a p53-independent manner, the exact molecular targets of EBR are still under investigation. Our recent SILAC (Stable Isotope Labeling by Amino Acids in Cell Culture) data showed that the most significantly altered protein after EBR treatment was calreticulin (CALR). CALR, a chaperone localized in endoplasmic reticulum (ER) lumen, plays role in protein folding and buffering Ca2+ ions. The alteration of CALR may cause ER stress and unfolded protein response correspondingly the induction of apoptosis. Unfolded proteins are conducted to 26S proteasomal degradation following ubiquitination. Our study revealed that EBR treatment caused ER stress and UPR by altering CALR expression causing caspase-dependent apoptosis in HCT 116, HT29, DLD-1, and SW480 colon cancer cells. Furthermore, 48 h EBR treatment did not caused UPR in Fetal Human Colon cells (FHC) and Mouse Embryonic Fibroblast cells (MEF). In addition our findings showed that HCT 116 colon cancer cells lacking Bax and Puma expression still undergo UPR and related apoptosis. CALR silencing and rapamycin co-treatment prevented EBR-induced UPR and apoptosis, whereas 26S proteasome inhibition further increased the effect of EBR in colon cancer cells. All these findings showed that EBR is an ER stress and apoptotic inducer in colon cancer cells without affecting nonmalignant cells.
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EBR induced autophagy in colon carcinoma cell lines
(2015-07) Adacan, Kaan; Çoker Gürkan, Ajda; Ünsal, Zeynep Narçin; ARISAN, ELİF DAMLA; YERLİKAYA, PINAR OBAKAN; 156421; 113920; 125860; 6125
Open Access
Endoplasmic reticulum stress is activated after epibrassinolide treatment leading autophagy in wild type and Atg5/ mouse embryonic fibroblasts: a survival mechanism in nonmalignant cells.
(2017) Adacan, Kaan; Çoker Gürkan, Ajda; Ünsal, Zeynep Narçin; ARISAN, ELİF DAMLA; YERLİKAYA, PINAR OBAKAN; 156421; 113920; 125860; 6125
Autophagy is a process of cellular selfdegradation during which macromolecules, damaged/aged organelles and proteins are delivered to the lysosome by engulfment within doublemembrane vesicles. The accumulation of un/misfolded proteins in the ER can activate ER stress leading unfolded protein response (UPR) resulting autophagy. Beclin1, LC3 and Autophagy Related Genes (Atg) play key roles in formation of autophagosome membrane, which are triggered by Ulk1 signaling through AMPK (AMPactivated protein kinase). Conversely, autophagy is inhibited by the mammalian target of rapamycin (mTOR), a central cellgrowth regulator that integrates growth factor and nutrient signals. Epibrassinolide (EBR) is a member of brassinostreoids plant hormones with a structural similarity to mammalian steroids. Our previous work demostrated that EBR treatment caused ER stress and UPR which resulted in caspasedependent apoptosis in SW480, DLD1, HT29 and HCT116 colon cancinoma cell lines. Here, we demostrate that EBR treatment activates UPR by inducing ER stressrelated proteins in wild type (wt) MEF (mouse embryonic fibroblast) cells. In contrary to our previous results in malignant cells, MEF cells do not undergo apoptosis after EBR treatment in stead autophagy is activated causing cell survival. We used Atg5/ MEF cells to understand the correlation between ER stress and autophagy. Suprisingly, we observed a significant downregulation of p62/SQSTM1 protein, which is known as one of the autophagy marker, in Atg5/ MEF cells. ER stress was also triggered in these cells. Therefore, we conclude that autophagy is a key mechanism to promote cell survival after EBR treatment against ER stress in both wt and Atg5/ MEF cells. These findings suggested that EBR as an ER stress inducer, is able to activate autophagy in order to promote cell survival in nonmalignant cells.
Open Access
Epibrassinolide Activates AKT to Trigger Autophagy With Polyamine Metabolism in SW480 and DLD-1 Colon Cancer Cell Lines
(The Scientific and Technological Research Council of Turkey, 2020) Adacan, Kaan; YERLİKAYA, PINAR OBAKAN
Epibrassinolide (EBR), a plant-derived polyhydroxylated derivative of 5 alpha-cholestane, structurally shows similarities to animal steroid hormones. According to the present study, EBR treatment triggered a significant stress response via activating ER stress, autophagy, and apoptosis in cancer cells. EBR could also increase Akt phosphorylation in vitro. While the activation of Akt resulted in cellular metabolic activation in normal cells to proceed with cell survival, a rapid stress response was induced in cancer cells to reduce survival. Therefore, Akt as a mediator of cellular survival and death decision pathways is a crucial target in cancer cells. In this study, we determined that EBR induces stress responses through activating Akt, which reduced the mTOR complex I (mTORC1) activation in SW480 and DLD-1 colon cancer cells. As a consequence, EBR triggered macroautophagy and led to lipidation of LC3 most efficiently in SW480 cells. The cotreatment of spermidine (Spd) with EBR increased lipidation of LC3 synergistically in both cell lines. We also found that EBR promoted polyamine catabolism in SW480 cells. The retention of polyamine biosynthesis was remarkable following EBR treatment. We suggested that EBR-mediated Akt activation might determine the downstream cellular stress responses to induce autophagy related to polyamines.
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Epibrassinolide controls polyamine levels to induce autophagy in Mouse Embryonic Fibroblasts
(2016) Adacan, Kaan; Çoker Gürkan, Ajda; Ünsal, Zeynep Narçin; ARISAN, ELİF DAMLA; YERLİKAYA, PINAR OBAKAN; 156421; 113920; 125860; 6125
Open Access
Epibrassinolide Treatment Caused Autophagy or Apoptosis Decision in a Time-Dependent Manner through ER Stress in Colon Cancer Cells
(2017) Adacan, Kaan; Çoker Gürkan, Ajda; Ünsal, Zeynep Narçin; ARISAN, ELİF DAMLA; YERLİKAYA, PINAR OBAKAN; 156421; 113920; 125860; 6125
Epibrassinolide (EBR) is a natural plant polyhydroxysteroid with structural similarity to steroid hormones. Lately we showed by SILAC assay that EBR treatment induces apoptosis by significantly altering the expressions of proteins having role in unfolded protein response (UPR) and endoplasmic reticulum (ER) stress. ER stress level has been proposed as a critical step in cancer chemotherapy. Moderate ER stress was shown as an inducer of pro-survival machinery via autophagy, which is an important catabolic process that delivers cytoplasmic material to the lysosome for degradation. However chronic ER stress lead cells to apoptosis. EBR treatment induced ER stress in a time-dependent manner within 48 h in SW480 and DLD1 colon cancer cells. Downregulation of p62, LC3 and increase in ATGs expressions indicated that autophagy is induced in these cell lines after 12 h EBR exposure. The mammalian target of rapamycin (mTOR), a coordinator between nutritional stress and cellular growth machinery, is associated with ER stress. Our results indicated that short time EBR exposure induced mTOR expression accompanied by Ser2448 dephosphorylation. After 48 h EBR treatment, with prolonged ER stress, both cell lines undergo apoptosis. Therefore we conclude that time-dependent EBR treatment caused autophagy or apoptosis decision through ER stress in colon cancer cells.
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Epibrassinolide-Induced Autophagy Occurs in an Atg5-Independent Manner Due to Endoplasmic Stress Induction in MEF Cells
(Springer, 2020) Adacan, Kaan; YERLİKAYA, PINAR OBAKAN; Çoker-Gürkan, Ajda; Kaya, Resul İsmail; Palavan-Ünsal, Narcin; ARISAN, ELİF DAMLA
Epibrassinolide (EBR), a polyhydroxysteroid belongs to plant growth regulator family, brassinosteroids and has been shown to have a similar chemical structure to mammalian steroid hormones. Our findings indicated that EBR could trigger apoptosis in cancer cells via induction of endoplasmic reticulum (ER) stress, caused by protein folding disturbance in the ER. Normal cells exhibited a remarkable resistance to EBR treatment and avoid from apoptotic cell death. The unfolded protein response clears un/misfolded proteins and restore ER functions. When stress is chronic, cells tend to die due to improper cellular functions. To understand the effect of EBR in non-malign cells, mouse embryonic fibroblast (MEF) cells were investigated in detail for ER stress biomarkers, autophagy, and polyamine metabolism in this study. Evolutionary conserved autophagy mechanism is a crucial cellular process to clean damaged organelles and protein aggregates through lysosome under the control of autophagy-related genes (ATGs). Cells tend to activate autophagy to promote cell survival under stress conditions. Polyamines are polycationic molecules playing a role in the homeostasis of important cellular events such as cell survival, growth, and, proliferation. The administration of PAs has been markedly extended the lifespan of various organisms via inducing autophagy and inhibiting oxidative stress. Our data indicated that ER stress is induced following EBR treatment in MEF cells as well as MEF Atg5(-/-) cells. In addition, autophagy is activated following EBR treatment by targeting PI3K/Akt/mTOR in wildtype (wt) cells. However, EBR-induced autophagy targets ULK1 in MEF cells lacking Atg5 expression. Besides, EBR treatment depleted the PA pool in MEF cells through the alterations of metabolic enzymes. The administration of Spd with EBR further increased autophagic vacuole formation. In conclusion, EBR is an anticancer drug candidate with selective cytotoxicity for cancer cells, in addition the induction of autophagy and PA metabolism are critical for responses of normal cells against EBR.
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Orlistat is a fatty acid synthase (FASN) inhibitor caused the modulation of AMPK and lipogenesis signaling axis in PC3 prostate cancer cells but not in PNT1A prostate epithelial cells
(2017) Nebiler, Esra; Adacan, Kaan; Çoker Gürkan, Ajda; Palavan Ünsan, Narçın; ARISAN, ELİF DAMLA; YERLİKAYA, PINAR OBAKAN; 113920; 125860; 156421; 6125
Prostate cancer is one the leading cancer type after lung cancer among male. Obesity is one of the known risk factor for developing highly aggressive and metastatic prostate cancer, which is an obstacle in cancer therapy. For these particular reasons, drugs are used in the therapy of obesity is suggested with their antitumoral effect, whether blocking adipogenesis and lipogenesis pathways or not. Orlistat inhibits fatty acid synthase (FASN), a large multifunctional enzyme, is responsible for de novo synthesis of long chain fatty acids. FASN activity is confirmed in the most of the xenograft models of prostate cancer cells. In this study we utilized PC3 cells, medium FASN expression profile compared to DU145 and LNCaP cells, and PNT1A prostate epithelial cells to evaluate the molecular targets of orlistat in adenosin mono phosphate kinase (AMPKa) and lipogenesis signaling axis. We demonstrated that orlistat inhibited cell proliferation and led to cell viability decrease in PC3 prostate cancer cells but not in PNT1A prostate epithelial cells. The activation of AMPKa was observed in both cell lines, which led to inhibition of protein translation through dephosphorylating mTOR at ser2448 and phosphorylating mTOR at ser2481 after treatment with Orlistat at 15 µM and 20 µM concentrations for 24h. Concomitantly, orlistat induced autophagy via phosphorylation of ULK1 at ser555. The modulation of AcetylCoA carboxylase (ACC), which regulates biosynthesis and oxidation of fatty acids was observed following orlistat treatment. However, AMPK phosphorylates ACC at ser79 and inhibits its activity. These findings suggest that orlistat altered AMPKrelated a number of critical pathways to exert its antitumor activity in prostate cancer cells. However, AMPK is a metabolic stress regulator protects epithelial normal cells against orlistat treatment. Thereby, it is crucial to have deep understanding about the role of AMPK in prostate cancer cells.