Effect Of FAM98A On Epithelial-mesenchymal Transition In Breast Cancer And Its Mechanism

Objective: The FAM98A(Family with Sequence Similarity 98,Member A)gene plays a role in many solid tumors,but its specific role has not been reported.There are currently relevant studies showing that it acts as a substrate for arginine methyltransferase(PRMT1)in ovarian cancer,PRMT1 can promote epithelial-mesenchymal transition(EMT)in ovarian cancer,The study found that FAM98 A as its substrate can also promote the colony formation of ovarian cancer cells,in vivo proliferation,migration and invasion.EMT is an important biological process for the ability of epithelial cell-derived malignant tumor cells to gain migration and invasion.However,the role of FAM98 A in tumorigenesis and development of breast cancer and its related regulatory mechanisms have rarely been reported.In this study,we first studied the effect of FAM98 A on the behavior of breast cancer cell proliferation,migration and invasion at the molecular level,and further study whether FAM98 A affect the behavior of breast cancer cells and its related molecular mechanisms by affecting EMT.Content: We detected the expression levels of FAM98 A in breast cancer cell lines and normal breast epithelial cells,and defined the effects of FAM98 A on breast cancer cell proliferation,migration,and invasion behaviors,and preliminary studied the molecular mechanism of FAM98 A regulating breast cancer cell EMT.Method: In this study,we first detected the expression level of FAM98 A protein in breast cancer cell lines and normal breast epithelial cells by Western Blot assay.We used the PCR experiment to massively amplify the target gene FAM98 A and construct the FAM98 A by linking it to the lentivirus system vector,so we constructed an overexpression plasmid for FAM98 A.We selected a cell line with a low expression level of the target gene and introduced the plasmid overexpressing FAM98 A into the stable line that overexpressed the target gene FAM98 A through a virus infection technique.At the same time,we selected a cell line with high expression of FAM98 A,and we selected the corresponding cell line.Interfering with the expression of FAM98 A in siRNA-introduced cells reduces the expression of FAM98 A.We used MTT,plate cloning,transwell,and scratch experiments to verify the effect of FAM98 A expression on cell traits such as breast cancer cell proliferation,migration,and invasion.We used Western Blot assay to detect the relationship between FAM98 A and EMT-related protein expression levels.Protein-profiling analysis and Coomassie Brilliant Blue staining technique were used to determine the relevant proteins regulated by FAM98 A in breast cancer,and the correlation between the relevant protein and FAM98 A was confirmed by CO-IP experiments in exogenous and endogenous systems,respectively.Our research can confirm the role of FAM98 A in breast cancer cell ethology and its related molecular mechanisms,providing a new target and molecular marker for the diagnosis and treatment of breast cancer.Results: We found that FAM98 A was highly expressed in ER-positive breast cancer cell lines(MCF-7,T47D)and low expression in ER-negative breast cancer cell lines(MDA-MB-231).We found that FAM98 A was normal High expression in mammary epithelial cells.Silencing its expression in FAM98 A overexpressing cell lines promotes the proliferation of breast cancer cells and inhibits the Invasion and metastasis capacity of breast cancer cells,in which the epithelial cell phenotype-associated protein: E-cadherin,Keratin expression is elevated,and the interstitial cell phenotype related protein N-cadherin and Vimentin expression decreased.We introduced FAM98 A plasmid into the FAM98 A low expression cell line to promote its expression to inhibit the proliferation of breast cancer cells and promote the invasion and metastasis of breast cancer cells,among which epithelial cell phenotype-associated protein: The expression of E-cadherin and keratin decreased,while the expression of N-cadherin and Vimentin related proteins in the stromal cell phenotype increased.To further explore the molecular mechanism of FAM98 A regulating EMT,we found through co-IP experiments that FAM98 A can bind directly to ER in exogenous 293 T cell line and endogenous MCF-7 cell line.We continued the Western Blot trial and found that when FAM98 A expression increased,the expression of ER was regulated,and the inhibitory effect of ER on downstream snail expression was relieved.Snail expression increased and inhibited the expression of downstream E-cadherin.This process eventually promoted breast cancer cell develop EMT.Conclusion: In breast cancer,the expression of FAM98 A is up-regulated.By binding to ER,it inhibits the expression of ER.Decreased expression of ER can relieve the inhibition of downstream snail expression.Increased expression of snail inhibits the expression of downstream E-cadherin.The process ultimately promotes EMT in breast cancer cells.