Molecular Mechanism Of NFIC1 Inhibiting The Migration And Invasion Of Breast Cancer Cells

Breast cancer is the most commonly diagnosed cancer in women.Due to the improvements in early detection and treatment,the 5-year survival rate of breast cancer patients has been increased gradually in recent years.However,the recurrence rate remains high in breast cancer patients,which is the leading cause of the disease-related mortality.The high recurrence rate is mainly caused by metastasis.Triple negative breast cancer(TNBC),a highly invasive subtype of breast cancer,has the highest metastasis rate.However,because of the highest incidence rate of luminal A breast cancer,the number of metastasis cases of luminal A breast cancer is higher than those of other breast cancer subtypes,although luminal A breast cancer patients generally have better prognosis.Many essential signaling transduction pathways that could promote metastasis are mis-regulated during tumorigenesis as a consequence of abnormal expression of key genes that are involved in these pathways.Therefore,identifying the key genes and pathways that could promote breast cancer metastasis is crucial for developing effective therapeutic approaches to prevent or treat recurrence,especially for TNBC and luminal A breast cancers.NFIC is a potent member in the family of nuclear factor I(NFI),which is capable of regulating mammary gland development and the expression of lactation related genes.There are five isoforms in NFIC.NFIC3 could inhibit the migration and invasion of breast cancer cells,and NFIC5 is a tumor suppressor in TNBC with anti-proliferative properties.NFIC1,the longest isoform of NFIC,is essential for the regulation on spatiotemporal expressions of drug-metabolizing genes in liver.However,the roles of NFIC1 in breast cancer metastasis and the underlying mechanisms have not been reported.Since the gene expression patterns and activated signaling transduction pathways are different between luminal A and triple negative breast cancer,the current study has explored the effects of NFIC1 on the migration and invasion of both luminal A and triple negative breast cancer cells.NFIC1 expression plasimid and si RNAs were transfected into MCF7 cells and MDA-MB-231 cells.Wound healing and Transwell assays showed that overexpression of NFIC1 suppressed the migration and invasion of MCF7 and MDA-MB-231 cells,and knockdown of NFIC1 enhanced the migration and invasion of MCF7 and MDA-MB-231 cells.To explore the mechanism by which NFIC1 inhibited the migration and invasion of breast cancer cells,RNA sequencing was used to identify the differentially expressed genes in MCF7 cells and MDA-MB-231 cells with or without NFIC1 overexpression.The RNA-seq analysis on MCF7 cells revealed that the interferon-activated Jak-STAT pathway was significantly enriched upon NFIC1 overexpression.Consistently,NFIC1 overexpression increased the expression and secretion of IFNB1,IFNL1,IFNL2 and IFNL3,and the activation of Jak-STAT pathway was enhanced by NFIC1 overexpression.In addition,treatment with Jak-STAT pathway inhibitors,Filgotinib or Ruxolitinib,reversed the suppressive effects of NFIC1 overexpression on migration and invasion of MCF7 cells.Furthermore,MX1,MX2 and RARRES3,the downstream target genes of Jak-STAT pathway,were knocked down in MCF7 cells with NFIC1 overexpression.MX1 and MX2 knockdown reversed the NFIC1-mediated inhibition on the migration and invasion of MCF7 cells.In addition,knockdown of IFNL1,IFNL2/3 and IFNB1 repressed NFIC1-induced Jak-STAT pathway,decreased expression of MX1 and MX2,and attenuated the inhibitory effects of NFIC1 on cell migration and invasion.To summarize,these results demonstrated that NFIC1 could enhance the expression of IFNL1,IFNL2/3 and IFNB1,which activates Jak-STAT pathway and subsequently inhibits the migration and invasion of MCF7 cells by stimulating the expressions of MX1 and MX2.In MDA-MB-231 cells,NFIC1 increased the transcription of S100A2 by binding to the promoter of S100A2.Knockdown of S100A2 reversed the suppressive effects of NFIC1 on cell migration and invasion.Besides,overexpression of NFIC1 reduced the phosphorylation of MEK and ERK,and knockdown of S100A2 reversed the suppressive effects of NFIC1 on MEK/ERK pathway.Treatment with the MEK/ERK pathway inhibitor,U0126,abolished the reversion effects mediated by S100A2 knockdown.In addition,MEK/ERK pathway was the downstream mediator of NFIC1-induced transition of MDA-MB-231 cells from mesenchymal properties to epithelial properties.Taken together,these results suggested that NFIC1 could inhibit the epithelial-mesenchymal transition of MDA-MB-231 cells by increasing S100A2-mediated inactivation of MEK/ERK pathway,which suppresses the migration and invasion of MDA-MB-231 cells.Collectively,our study investigated the inhibitory effects of NFIC1 on the migration and invasion of MCF7 cells and MDA-MB-231 cells,and revealed the molecular mechanism underlying NFIC1-mediated inhibition of migration and invasion on both MCF7 and MDA-MB-231 cells.NFIC1 suppresses migration and invasion of MCF7 cells through interferon-mediated Jak-STAT pathway,and suppresses the migration and invasion of MDA-MB-231 cells through increasing S100A2-induced inactivation of MEK/ERK pathway.Our study provides a new discovery on the mechanism of NFIC1-mediated inhibition of the migration and invasion of breast cancer cells,and suggests that NFIC1 might be a potential target for the treatment of breast cancer metastasis.