Browsing by Author "Alkurt, Gizem"

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    PublicationOpen Access
    Co-Chaperone Bag-1 Plays a Role in the Autophagy-Dependent Cell Survival Through Beclin 1 Interaction
    (MDPI, 2021) Türk, Miray; Tatlı, Özge; Alkan, Hamza Furkan; KILBAŞ, PELİN ÖZFİLİZ; Alkurt, Gizem; Dinler Doğanay, Gizem
    Expression levels of the major mammalian autophagy regulator Beclin 1 and its interaction with Bcl-2 regulate the switch between autophagic cell survival and apoptotic cell death pathways. However, some of the regulators and the precise mechanisms of these processes still remain elusive. Bag-1 (Bcl-2 associated athanogene-1), a member of BAG family proteins, is a multifunctional pro-survival molecule that possesses critical functions in vital cellular pathways. Herein, we report the role of Bag-1 on Bcl-2/Beclin 1 crosstalk through indirectly interacting with Beclin 1. Pull-down experiments suggested a molecular interaction between Bag-1 and Beclin 1 in breast cancer cell lines. On the other hand, in vitro binding assays showed that Bag-1/Beclin 1 interaction does not occur directly but occurs through a mediator molecule. Bag-1 interaction with p-Beclin 1 (T119), indicator of early autophagy, is increased during nutrient starvation suggesting involvement of Bag-1 in the autophagic regulation. Furthermore, CRISPR/Cas9-mediated Bag-1 knock-out in MCF-7 cells hampered cell survival and proliferation and resulted in decreased levels of total LC3 under starvation. Collectively, we suggest that Bag-1 modulates cell survival/death decision through maintaining macroautophagy as a component of Beclin 1-associated complexes.
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    PublicationOpen Access
    The Comparison of Differentially Expressed microRNAs in Bag-1 Deficient and Wild Type MCF-7 Breast Cancer Cells by Small RNA Sequencing
    (TUBITAK Scientific & Technical Research Council Turkey, 2022) KILBAŞ, PELİN ÖZFİLİZ; Alkurt, Gizem; Çoker Gürkan, Ajda; DİNLER DOĞANAY, GİZEM; ARISAN, ELİF DAMLA; YERLİKAYA, PINAR OBAKAN
    The multifunctional BAG-1 (Bcl-2 athanogene-1) protein promotes breast cancer survival through direct or indirect interaction partners. The number of the interacting partners determines its cellular role in different conditions. As well as interaction partner variability, the amount of BAG-1 protein in the cells could cause dramatic alterations. According to previous studies, while the transient silencing of Bag-1 enhanced drug-induced apoptosis, deletion of BAG-1 could induce stemness properties and Akt-mediated actin remodeling in MCF-7 breast cancer cells. Considering the heterogeneity of breast cancer and the variability of BAG-1-mediated cell response, it has become essential to determine microRNA (miRNA) functions in breast cancer depending on Bag-1 expression level. This study aims to compare microRNA expression levels in wt and Bag-1 knockout (KO) MCF-7 breast cancer cells. hsa-miR-429 was selected as a potential miRNA in BAG-1KO MCF-7 cells because of the downregulation both in bioinformatics and validation qRT-PCR assay. According to predicted mRNA targets and functional enrichment analysis the ten hub proteins that are phosphatidylinositol4,5-biphosphate 3-kinase catalytic subunit alpha (PIK3CA), kinase insert domain receptor (KDR), GRB2 associated binding protein 1 (GAB1), Rac family small GTPase1 (RAC1), vascular endothelial growth factor A (VEGFA), Cbl proto-oncogene (CBL), syndecan 2 (SDC2), phospholipase C gamma 1 (PLCG1), E1A binding protein p300 (EP300), and CRK like proto-oncogene, adaptor protein (CRKL) were identified as targets of hsa-miR-429. The functional enrichment analysis showed that the most significant proteins were enriched in PI3K/Akt, focal adhesion, cytoskeleton regulation, proteoglycans in cancer, and Ras signaling pathways. It was determined that hsa-miR-429 targeted these pathways in Bag-1 deficient conditions and could be used as a potential therapeutic target in future studies.
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    PublicationEmbargo
    Downregulation of c-Myc mediated ODC expression after purvalanol treatment is under control of upstream MAPK signaling axis in MCF-7 breast cancer cells
    (Tubitak Scientific & Technical Research Council Turkey, Ataturk Bulvarı No 221, Kavaklıdere, Ankara, 00000, Turkey, 2014) Alkurt, Gizem; Köse, Betsi; Çoker Gürkan, Ajda; Palavan Unsal, Narçin; ARISAN, ELİF DAMLA; YERLİKAYA, PINAR OBAKAN; COŞKUN, DENİZ; 156421; 125860; 113920; 6125
    Roscovitine and purvalanol are specific cyclin-dependent kinase (CDK) inhibitors, which induce apoptosis by triggering cell cycle arrest in various cancer cells such as colon, prostate, and breast cancer cells. Although the apoptotic action of roscovitine was clarified at the molecular level, the exact mechanism of purvalanol-induced apoptosis is still under investigation. The mitogen-activated protein kinase (MAPK) signaling cascade is activated by different inducers related to growth, proliferation, differentiation processes, or environmental stress factors. Recent reports showed that modulation of MAPKs might lead to regulation of c-Myc, which is a transcription factor for the polyamine (PA) biosynthesis enzyme, ornithine decarboxylase (ODC). PAs are amine-derived cationic molecules that play crucial roles in cell proliferation, growth, and differentiation. In this study, we investigated the potential role of the MAPK signaling cascade in the purvalanol-induced apoptosis mechanism by comparing the results of roscovitine in MCF7 and MDA-MB-231 breast cancer cells. We found that CDK inhibitors decreased the cell viability in a dose-and time-dependent manner in MCF-7 and MDA-MB-231 cancer cells. Although both CDK inhibitors induced cell cycle arrest, which led to apoptosis by activating caspases and PARP cleavage in MCF-7 breast cancer cells, the apoptotic effect of purvalanol was less than that of roscovitine in MDA-MB-231 cells. Inhibition of MAPKs prevented CDK inhibitor-induced cell viability loss in both cell lines. We determined that purvalanol downregulated c-Myc and ODC expression levels, which led to sharp decrease in the PA pool in MCF-7 cells. On the contrary, purvalanol did not significantly alter c-Myc expression levels, which led to de novo biosynthesis of ODC in a time-dependent manner in MDA-MB-231 cells. Therefore, we suggest that a purvalanol-mediated resistance phenotype might be a possible outcome of c-Myc-mediated ODC expression level in MDA-MB-231 cells.
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    ODC is a Mediator of the Purvalanol A-Induced Apoptotic Signaling via the p38 Mitogen-Activated Protein Kinase Pathway in MCF-7 Breast Cancer Cells
    (2012) Köse, Betsi; Alkurt, Gizem; Çoker Gürkan, Ajda; Ünsal, Zeynep Narçin; ARISAN, ELİF DAMLA; 125860; 113920; 6125