The effect of SOX۲ overexpression on OCT۴and LGR۵ genes transcription and SOX۲ autoregulation insw۴۸۰ cell line

Background: Colorectal cancer (CRC), the third deadliest cancer,has gotten the attention of many researchers. In most cases,a loss-of-function mutation in tumor suppressor genes like APCbrings about neoplasia in the epithelium tissue of the colon andthe rectum, resulting in the formation of a primary tumor. As thecells proliferate, other cancerous mutations accumulate in theirgenome, facilitating adenoma to carcinoma transition. It hasshown that in multiple cancers, including glioblastoma, breast,and ovarian, SOX۲ level increases during cancer progressionand affects cell physiology. It has also shown that the relationshipbetween SOX۲ and specific transcription factors, includingOCT۴ and autoregulation of SOX۲, can induce a stem-like statein cancer cells. In a normal colon crypt LGR۵ surface receptoris a hallmark of stem cells which has demonstrated upregulationin stem-like cells inside the tumor tissue. These cells areassumed to participate in many pathological malignancy conditions,such as drug resistance, recurrence, and metastasis. Accordingto the importance of the SOX۲ protein, the relationshipbetween SOX۲, OCT۴, and LGR۵ in CRC cell line SW۴۸۰ hasbeen investigated in the present study.Materials and Methods: SOX۲ gene was sub-cloned frompSin-EF۱a-SOX۲-Pur into the pIRES-DsRED-Express۲ plasmid.SW۴۸۰ cells were seeded into a ۲۴-well plate and treatedwith ۲% DMSO before transfection. Transfection of sw۴۸۰using pIRES-DsRED-SOX۲-Express۲ and pIRES-DsRES-Express as mock vector conducted with Lipofectamine ۲۰۰۰. Forproducing stable transfectants, the G۴۱۸ antibiotic was used fortwo weeks. The expression level of mentioned factors was thenassessed by real-time PCR.Results: Twenty-four hours and ۱۴ days after transfection,the fluorescent imaging showed red fluorescence in mock andSOX۲ transfectants. According to qRT-PCR data, the totalSOX۲ mRNA was elevated (۵.۸-fold), OCT۴ transcript wasincreased (۱۵-fold), endogenous expression of SOX۲ was upregulated(۲.۹-fold), and LGR۵ level was slightly changed (۱.۴-fold) in comparison with mock cells.Conclusion: In this study, induction of OCT۴ and LGR۵, andSOX۲ autoregulation by ectopic expression of SOX۲ was observed,which is also demonstrated in other cancer cell types.This change in the expression of stemness markers may, in turn,affect cancer cell characteristics.