Moleküler Biyoloji ve Genetik Bölümü / Department of Molecular Biology and Genetics
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Publication 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 OBAKANThe 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.Publication CRISPR/Cas9-Mediated Bag-1 Knockout Increased Mesenchymal Characteristics of MCF-7 Cells Via Akt Hyperactivation-Mediated Actin Cytoskeleton Remodeling(Public Library of Science, 2022) KILBAŞ, PELİN ÖZFİLİZ; Can, Nisan Denizce; Kızılboğa, Tuğba; Ezberci, Fikret; Doğanay, Hamdi Levent; Doğanay, Gizem Dinler; ARISAN, ELİF DAMLABag-1 protein is a crucial target in cancer to increase the survival and proliferation of cells. The Bag-1 expression is significantly upregulated in primary and metastatic cancer patients compared to normal breast tissue. Overexpression of Bag-1 decreases the efficiency of conventional chemotherapeutic drugs, whereas Bag-1 silencing enhances the apoptotic efficiency of therapeutics, mostly in hormone-positive breast cancer subtypes. In this study, we generated stable Bag-1 knockout (KO) MCF-7 breast cancer cells to monitor stress-mediated cellular alterations in comparison to wild type (wt) and Bag-1 overexpressing (Bag-1 OE) MCF-7 cells. Validation and characterization studies of Bag-1 KO cells showed different cellular morphology with hyperactive Akt signaling, which caused stress-mediated actin reorganization, focal adhesion decrease and led to mesenchymal characteristics in MCF-7 cells. A potent Akt inhibitor, MK-2206, suppressed mesenchymal transition in Bag-1 KO cells. Similar results were obtained following the recovery of Bag-1 isoforms (Bag-1S, M, or L) in Bag-1 KO cells. The findings of this study emphasized that Bag-1 is a mediator of actin-mediated cytoskeleton organization through regulating Akt activation. © 2022 Kilbas et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.Publication The Protective Impact of Growth Hormone Against Rotenone-Induced Apoptotic Cell Death via Acting on Endoplasmic Reticulum Stress and Autophagy Axis(Scientific and Technological Research Council Turkey, 2023) RENCÜZOĞULLARI, ÖZGE; TORNACI, SELAY; ÇELİK, YAĞMUR; TAŞ, NAYAT NAROT; YERLİKAYA, PINAR OBAKAN; Arısan, Elif Damla; Gürkan, Ajda ÇokerHuman growth hormone (GH) is crucial modulator of cellular metabolisms, including cell proliferation and organ development, by stimulating insulin-like growth factor-1 (IGF-1), which has various functions such as cell proliferation, tissue growth, survival, or neuroprotection. Therefore, GH is implicated as a critical player in the cell and can enhance neurogenesis and provide neuroprotection during the treatment of neurological diseases such as Parkinson's disease (PD). In this study, the neuroprotective role of GH was investigated in rotenone-induced PD models for the first time. Both SH-SY5Y and SK-N-AS neuroblastoma cells were exposed to rotenone to mimic PD pathogenesis as stated in previous studies. Our data demonstrated that overexpression of GH led to the resistance of the SH-SY5Y and SK-N-AS cell lines to rotenone treatment. The levels of ER stress markers, CHOP, PERK, XBP-1, and ATF6, were higher in wt cells than GH+ SH-SY5Y cells. However, the level of autophagy markers LC3 increased and the levels of reactive oxygen species (ROS) decreased with the overexpression of GH. Furthermore, while rotenone significantly increased the SubG1 population in the cell cycle of SH-SY5Y wt cells, there was a minor alteration in GH+ cell population. Concomitantly, the levels of the proapoptotic marker, cleaved-PARP, and positive staining of Annexin V in SH-SY5Y wt cells were higher after rotenone treatment. Together, these results revealed that overexpression of GH enhanced the autophagy response by triggering the ER stress of SH-SY5Y cells to rotenone exposure and showed a neuroprotective effect in vitro PD models.