ELF3 Expression Research Articles

CDK5 Upregulated by ELF3 Transcription Promotes IL-1β-induced Inflammation and Extracellular Matrix Degradation in Human Chondrocytes.

Osteoarthritis (OA) is a common chronic disease with age-associated increase in both incidence and prevalence. The cyclin-dependent kinase 5 (CDK5), which is a member of the CDK family, is involved in many chronic diseases. This study was performed to explore the functional role of CDK5 in OA and to discuss the detailed molecular mechanisms. The expressions of CDK5 and ELF3 before or after transfection were detected with reverse transcription-quantitative PCR (RT-qPCR) and western blot. 5-ethynyl-2'-deoxyuridine (Edu) and terminal deoxynucleoitidyl transferase-mediated nick-end labeling (TUNEL) assays were used to detect the proliferation and apoptosis of C28/I2 cells. The levels of inflammatory cytokines were estimated using enzyme-linked immunosorbent assay (ELISA) while the expressions of proteins implicated in extracellular matrix (ECM) degradation- and apoptosis were detected using western blot. Additionally, the activity of CDK5 promoters and its binding with ELF3 were detected using luciferase activity assay and chromatin immunoprecipitation (CHIP) assay. In the present study, it was discovered that the mRNA and protein expressions of CDK5 were significantly increased in IL-1β-induced C28/I2 cells. After depleting CDK5 expression, the apoptosis, inflammation and ECM in C28/I2 cells with IL-1β induction were suppressed. It was also found that ELF3 expression was increased in IL-1β-induced C28/I2 cells and acted as a transcription factor binding to the CDK5 promoter to regulate its transcriptional expression. The further experiments evidenced that ELF3 overexpression partially reversed the inhibitory effects of CDK5 deficiency on IL-1β-induced apoptosis, inflammation and ECM in C28/I2 cells. Collectively, CDK5 that upregulated by ELF3 transcription could promote the development of OA.

TNBC response to paclitaxel phenocopies interferon response which reveals cell cycle-associated resistance mechanisms.

Paclitaxel is a standard of care neoadjuvant therapy for patients with triple negative breast cancer (TNBC); however, it shows limited benefit for locally advanced or metastatic disease. Here we used a coordinated experimental-computational approach to explore the influence of paclitaxel on the cellular and molecular responses of TNBC cells. We found that escalating doses of paclitaxel resulted in multinucleation, promotion of senescence, and initiation of DNA damage induced apoptosis. Single-cell RNA sequencing (scRNA-seq) of TNBC cells after paclitaxel treatment revealed upregulation of innate immune programs canonically associated with interferon response and downregulation of cell cycle progression programs. Systematic exploration of transcriptional responses to paclitaxel and cancer-associated microenvironmental factors revealed common gene programs induced by paclitaxel, IFNB, and IFNG. Transcription factor (TF) enrichment analysis identified 13 TFs that were both enriched based on activity of downstream targets and also significantly upregulated after paclitaxel treatment. Functional assessment with siRNA knockdown confirmed that the TFs FOSL1, NFE2L2 and ELF3 mediate cellular proliferation and also regulate nuclear structure. We further explored the influence of these TFs on paclitaxel-induced cell cycle behavior via live cell imaging, which revealed altered progression rates through G1, S/G2 and M phases. We found that ELF3 knockdown synergized with paclitaxel treatment to lock cells in a G1 state and prevent cell cycle progression. Analysis of publicly available breast cancer patient data showed that high ELF3 expression was associated with poor prognosis and enrichment programs associated with cell cycle progression. Together these analyses disentangle the diverse aspects of paclitaxel response and identify ELF3 upregulation as a putative biomarker of paclitaxel resistance in TNBC.

ELF3 plays an important role in defining TRAIL sensitivity in breast cancer by modulating the expression of decoy receptor 2 (DCR2).

TRAIL protein on binding to its cognate death receptors (DR) can induce apoptosis specifically in breast tumor cells sparing normal cells. However, TRAIL also binds to decoy receptors (DCR) thereby inhibiting the apoptotic pathways thus causing TRAIL resistance. Also, one of the barriers due to which TRAIL-based therapy could not become FDA-approved might be because of resistance to therapy. Therefore, in the current study we wanted to explore the role of transcription factors in TRAIL resistance with respect to breast cancer. Microarray data from TRAIL-sensitive (TS) and TRAIL-resistant (TR) MDA-MB-231 cells were reanalyzed followed by validation of the candidate genes using quantitative PCR (qPCR), immunoblotting and immunofluorescence technique. Overexpression of the candidate gene was performed in MDA-MB-231 and MCF7 cells followed by cell viability assay and immunoblotting for cleaved caspase-3. Additionally, immunoblotting for DCR2 was carried out. TCGA breast cancer patient survival was used for Kaplan-Meier (KM) plot. Validation of the candidate gene i.e. ELF3 using qPCR and immunoblotting revealed it to be downregulated in TR cells compared to TS cells. ELF3 overexpression in MDA-MB-231 and MCF7 cells caused reversal of TRAIL resistance as observed using cell viability assay and cleaved caspase-3 immunoblotting. ELF3 overexpression also resulted in DCR2 downregulation in the MDA-MB-231 and MCF7 cells. Furthermore, KM analysis found high ELF3 and low DCR2 expression to show better patient survival in the presence of TRAIL. Our study shows ELF3 to be an important factor that can influence TRAIL-mediated apoptosis in breast cancer. Also, ELF3 and DCR2 expression status should be taken into consideration while designing strategies for successful TRAIL-based therapy.

Progesterone-mediated remodeling of the maternal-fetal interface by a PGRMC1-dependent mechanism

Implantation and maintenance of pregnancy involve intricate immunological processes that enable the developing fetus to coexist with the maternal immune system. Progesterone, a critical hormone during pregnancy, is known to promote immune tolerance and prevent preterm labor. However, the mechanism by which progesterone mediates these effects remains unclear. In this study, we investigated the role of the non-classical progesterone receptor membrane component 1 (PGRMC1) in progesterone signaling at the maternal-fetal interface. Using JEG3 cells, a trophoblast model cell line, we observed that progesterone stimulation increased the expression of human leukocyte antigen-C (HLA-C) and HLA-G, key molecules involved in immune tolerance. We also found that progesterone upregulated the expression of the transcription factor ELF3, which is known to regulate trophoblast-specific HLA-C expression. Interestingly, JEG3 cells lacked expression of classical progesterone receptors (PRs) but exhibited high expression of PGRMC1, a finding we confirmed in primary trophoblasts by mining sc-RNA seq data from human placenta. To investigate the role of PGRMC1 in progesterone signaling, we used CRISPR/Cas9 technology to knockout PGRMC1 in JEG3 cells. PGRMC1-deficient cells showed a diminished response to progesterone stimulation. Furthermore, we found that the progesterone antagonist RU486 inhibited ELF3 expression in a PGRMC1-dependent manner, suggesting that RU486 acts as a progesterone antagonist by competing for receptor binding. Additionally, we found that RU486 inhibited cell invasion, an important process for successful pregnancy, and this inhibitory effect was dependent on PGRMC1. Our findings highlight the crucial role of PGRMC1 in mediating the immunoregulatory effects of progesterone at the maternal-fetal interface.

Exploring Gene Regulatory Networks and Potential Therapeutic Drugs Based on ELF3 Expression in Cholangiocarcinoma

Exploring Gene Regulatory Networks and Potential Therapeutic Drugs Based on ELF3 Expression in Cholangiocarcinoma

Comprehensive analysis of E47‑like factors and verification of ELF4 in clear cell renal cell carcinoma.

Clear cell renal cell carcinoma (ccRCC) is the most prominent subtype of renal cancer and E47-like factors (ELFs) are important in tumorigenesis; however, the specific role of key ELFs in ccRCC remains unclear. The present study comprehensively analyzed RNA sequencing and clinical data from multiple databases, and identified differentially expressed ELFs (ELF3-5) in ccRCC. The DNA promoter methylation, genetic variation and clinical significance of ELF3-5 in ccRCC were analyzed using the cBioPortal and UALCAN databases. The association between ELF3-5 and multiple immune cell infiltration was analyzed using Tumor Immune Estimation Resource. Subsequently, ELF4 was selected and its association with biological functions was assessed. Cell counting kit-8 (CCK-8), colony formation, Transwell, macrophage chemotaxis and polarization assays were conducted to validate the functions of ELF4. Notably, the mRNA expression levels of ELF4 were significantly upregulated in ccRCC, whereas ELF3 and ELF5 mRNA expression levels were significantly downregulated. Clinical significance analysis revealed that ELF4 showed a high clinical significance with tumor grade, clear cell type A and B subtypes, and incidence rates of amplification in genetic variation. Further analyses indicated that ELF4 may be involved in multiple immune cell differentiation. Additionally, cell experiments revealed that ELF4 inhibition downregulated 769-P and 786-O proliferation, migration and invasion. Knockdown of ELF4 in cancer cells also inhibited M2 macrophage polarization and chemotaxis towards 769-P and 786-O cells. Conclusively, the present findings indicated the clinical significance of ELF4 in ccRCC, and verified its key role in driving cell proliferation, migration and invasion, and promoting M2 macrophage polarization and chemotaxis in ccRCC.

Abstract 3894: The role of ELF3 in acquired resistance to endocrine therapy in ER-positive breast cancer

Abstract Endocrine therapies targeting the estrogen receptor (ER) are the major treatments of ER+ breast cancer, however acquired resistance to endocrine therapy is almost unavoidable and is the main cause of death with breast cancer. The only FDA-approved selective estrogen receptor degrader (SERD) fulvestrant has been used in the second line treatment to overcome aromatase inhibitor- or tamoxifen-resistance. However, fulvestrant has poor pharmacokinetic properties, which has inspired the development of a new generation of oral SERDs including amcenestrant and giredestrant. To understand the mechanisms of acquired resistance to SERDs in ER+ breast cancer, we established ER+ breast cancer cell lines (MCF7 and T47D) resistant to fulvestrant, amcenestrant or giredestrant. We found that all of our MCF7 and T47D SERD-resistant cell lines showed cross-resistance to each of the three SERDs. We then performed multiple genome-wide profiling assays including CRISPR-KO screen, whole exome sequencing (WES), RNA-seq, ATAC-seq and H3K27ac ChIP-seq on the parental and SERD resistant cell lines. Our findings from the CRISPR-KO screens suggest that ETS transcription factor ELF3 is a potential driver of resistance to SERDs. ELF3 expression is negatively regulated by estrogen signaling. Cells that developed acquired resistance to different SERDs consistently show depletion of ER expression but also a dramatic increase of ELF3 expression, while knockdown of ELF3 in the resistant cells leads to cell growth inhibition, suggesting that ELF3 can drive cell survival in the SERD resistant cells. When comparing the open chromatin regions and active enhancer regions between SERD resistant and parental cells via ATAC-seq and H3K27ac ChIP-seq respectively, ELF3 DNA binding motif was found significantly enriched in SERD resistant cells versus parental cells further indicating that ELF3 may play an important role in the SERD resistant cells. ELF3 ChIP-seq was also performed to identify genes enriched with ELF3 binding sites in SERD resistant and parental cells. Pathway analysis of ELF3-targeted genes reveals that ELF3 may regulate cell metabolism specifically in SERD resistant cells but not in parental cells. Our study suggests that ELF3 may be a novel driver of acquired resistance to SERDs in ER+ breast cancer. Citation Format: Na Zhang, Rongbin Zheng, Myles Brown. The role of ELF3 in acquired resistance to endocrine therapy in ER-positive breast cancer. [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 3894.

ELF3 promotes gemcitabine resistance through PKMYT1/CDK1 signaling pathway in gallbladder cancer.

Gemcitabine is the standard treatment for gallbladder cancer (GBC) patients, and the development of resistance frequently limits its efficacy. However, the molecular features and mechanisms of gemcitabine resistance (Gem-R) in GBC cells remain unknown. Herein, we aimed to explore the role of ELF3 in Gem-R of GBC, including the underlying mechanisms. RNA sequencing was used to screen the essential genes related to the generation of Gem-R in GBC tissues. The correlation between Gem-R and ELF3 expression was identified in GDSC, GEO database, GBC tissues, and 3 GBC cell lines. Immunohistochemical staining, quantitative real-time polymerase chain reaction, and western blot were used to examine the expression of ELF3, PKMYT1, and CDK1. Luciferase reporter assays were used to identify the binding site of ELF3 in the PKMYT1 promoter region. CCK-8 assay and clonogenic survival assays were used to evaluate the sensitivity of gemcitabine in GBC cells. A GBC xenograft model was used to evaluate the influence of ELF3 on the therapeutic effect of gemcitabine. A consistently positive correlation between ELF3 expression and Gem-R, both in newly generated GBC RNA-seq data and in the datasets from GDSC and GEO. Gem-R in GBC cells was facilitated by ELF3 overexpression, whereas ELF3 knockdown had the opposite effect. In vivo experiments further proved that reducing ELF3 expression promoted the gemcitabine sensitivity of GBC cells and extended the survival time of mice that received orthotopic xenografted tumors. Mechanistically, ELF3 upregulated PKMYT1 expression by interacting with the DNA binding region of PKMYT1 in GBC cells, thereby promoting the phosphorylation of CDK1 and inducing Gem-R. Treatment with a combination of the PKMYT1 shRNA and gemcitabine significantly reduced the growth of GBC cells induced by overexpression of ELF3 in vitro and in vivo. ELF3/PKMYT1/CDK1 axis significantly regulates Gem-R to GBC cells and may represent a promising drug target for treating GBC patients.

The transcription factor ELF5 is essential for early preimplantation development.

During early embryonic development, the cell adhesion molecule E-cadherin encoded by the Cdh1 gene plays a vital role in providing proper cell-cell adhesion, ensuring an undifferentiated state critical for maintaining the pluripotency for the development of the preimplantation embryo. The transcriptional regulation of Cdh1 gained attention recently but is not yet fully understood. In a previous study, our team established a correlation between Elf3 and Cdh1 expression and showed its importance in the regulation of MET. Here, the regulation of Cdh1 by Ets transcription factors in early embryogenesis was investigated. A loss-of-function approach was used to study the effect of Elf5 loss on Cdh1 gene expression by small interfering RNAs in fertilized oocytes. Changes in gene expression were measured by qPCR analysis, and developing embryos were visualized by microscopy. Loss of Elf5 arrested the embryos at the 2-cell stage, accompanied by a significant downregulation of Cdh1 expression. The findings presented here illustrate the role of ELF5 in preimplantation development and in regulating the expression of Cdh1. The maintenance of the ELF5 and Cdh1 regulatory node proved essential for the proper development of the early mouse embryos, which is in agreement with the critical role of Elf5 and Cdh1 genes in regulating the early events during embryogenesis.

LncRNA ELF3-AS1 inhibits gastric cancer by forming a negative feedback loop with SNAI2 and regulates ELF3 mRNA stability via interacting with ILF2/ILF3 complex

BackgroundThe biological function of lncRNA ELF3-AS1 remains largely unknown in cancers. The cause of SNAI2 overexpression in tumor metastasis remains largely unclear. The molecular mechanisms underlying the high co-expression of antisense lncRNAs and adjacent protein-coding genes remains unclear.MethodsRNA-seq, CHIP and dual-luciferase reporter assay were performed to identify lncRNAs regulated by SNAI2. MicroRNA-seq and RNA-seq studies were conducted to reveal the biological function of ELF3-AS1 in GC. RNA pulldown and CHIRP assays were conducted to identify the protein that interacts with ELF3-AS1.ResultsA total of 123 lncRNAs were identified to be regulated by SNAI2 in GC by RNA sequencing. The ELF3 gene and antisense lncRNA ELF3-AS1 were both transcriptionally repressed by SNAI2 or SNAI1. Down-regulation of ELF3-AS1 and ELF3 predicted poor prognosis in GC. Nuclear localized lncRNA ELF3-AS1 negatively regulated GC cell cycle progression via suppressing G1/S transition and histone synthesis. ELF3-AS1 mainly inhibited GC metastasis by repressing SNAI2 signaling. Additionally, ELF3-AS1 modulated ELF3 mRNA stability by RNA-RNA interaction. The RNA duplexes formed by ELF3 mRNA and lncRNA ELF3-AS1 directly interacted with the double-stranded RNA (dsRNA) binding protein complex ILF2/ILF3 (NF45/NF90). In turn, the ILF2/ILF3 complex dynamically regulated the expression of ELF3-AS1 and ELF3 by affecting the dsRNA stability.ConclusionsThe SNAI2-ELF3-AS1 feedback loop regulates ELF3 expression at transcriptional and post-transcriptional levels and drives gastric cancer metastasis by maintaining SNAI2 overexpression. The ILF2/ILF3 complex plays a critical role in regulating dsRNA stability. In addition, our work provides a direct evidence that head-to-head antisense lncRNAs can share promoters with neighboring coding genes, which make their expression subject to similar transcriptional regulation, leading to high co-expression.

Elf3 deficiency during zebrafish development alters extracellular matrix organization and disrupts tissue morphogenesis.

E26 transformation specific (ETS) family transcription factors are expressed during embryogenesis and are involved in various cellular processes such as proliferation, migration, differentiation, angiogenesis, apoptosis, and survival of cellular lineages to ensure appropriate development. Dysregulated expression of many of the ETS family members is detected in different cancers. The human ELF3, a member of the ETS family of transcription factors, plays a role in the induction and progression of human cancers is well studied. However, little is known about the role of ELF3 in early development. Here, the zebrafish elf3 was cloned, and its expression was analyzed during zebrafish development. Zebrafish elf3 is maternally deposited. At different developmental stages, elf3 expression was detected in different tissue, mainly neural tissues, endoderm-derived tissues, cartilage, heart, pronephric duct, blood vessels, and notochord. The expression levels were high at the tissue boundaries. Elf3 loss-of-function consequences were examined by using translation blocking antisense morpholino oligonucleotides, and effects were validated using CRISPR/Cas9 knockdown. Elf3-knockdown produced short and bent larvae with notochord, craniofacial cartilage, and fin defects. The extracellular matrix (ECM) in the fin and notochord was disorganized. Neural defects were also observed. Optic nerve fasciculation (bundling) and arborization in the optic tectum were defective in Elf3-morphants, and fragmentation of spinal motor neurons were evident. Dysregulation of genes encoding ECM proteins and matrix metalloprotease (MMP) and disorganization of ECM may play a role in the observed defects in Elf3 morphants. We conclude that zebrafish Elf3 is required for epidermal, mesenchymal, and neural tissue development.

E74 Like ETS Transcription Factor 3 is a Negative Regulator of Pathogenic Lamina Propria T Helper 17.1 Cells in Murine Colitis

ABSTRACT Interleukin-17A (IL-17A)-expressing T cells, including T helper 17 (Th17) and T helper 17.1 (Th17.1) cells, play a significant role in inflammatory bowel diseases (IBDs). Identifying the mechanisms underlying the heterogeneity and plasticity of IL-17A-expressing T cells is crucial for understanding and controlling their pathogenicity. The role of E74 like ETS transcription factor 3 (ELF3) in regulating the pathogenicity of IL-17A-expressing T cells has not been studied before. Dextran sulfate sodium was used to induce acute colitis in transgenic mice co-expressing IL-17A and enhanced green fluorescent protein (EGFP). IL-17A-expressing T cells were analyzed by flow cytometry. ELF3 expression was evaluated by reverse transcription and quantitative polymerase chain reaction. Lentivirus-mediated ELF3 overexpression was performed to assess the effect of ELF3 on Th17 and Th17.1 cells in vitro. The in vivo effect of ELF3 on Th17.1 cells was analyzed in an adoptive transfer colitis model. ELF3 was expressed by IL-17A-expressing T cells in the colonic lamina propria after colitis induction. Th17 cells and Th17.1 cells were distinguished based on the expression of C-X-C motif chemokine receptor 3, cytokine production, and key regulators. Th17 cells expressed higher ELF3 than Th17.1 cells. Ectopic ELF3 overexpression did not alter Th17 cell function while suppressing Th17.1 cell function in vitro. When adoptively transferred into Rag1 knockout mice to induce colitis, ELF3-overexpressing Th17.1 cells were less pathogenic than the control Th17.1 cells. ELF3 suppresses the pathogenicity of Th17.1 cells in colitis.

ETS transcription factor ELF3 (ESE-1) is a cell cycle regulator in benign and malignant prostate.

This study aimed to elucidate the role of ELF3, an ETS family member in normal prostate growth and prostate cancer. Silencing ELF3 in both benign prostate (BPH‐1) and prostate cancer (PC3) cell lines resulted in decreased colony‐forming ability, inhibition of cell migration and reduced cell viability due to cell cycle arrest, establishing ELF3 as a cell cycle regulator. Increased ELF3 expression in more advanced prostate tumours was shown by immunostaining of tissue microarrays and from analysis of gene expression and genetic alteration studies. This study indicates that ELF3 functions not only as a part of normal prostate epithelial growth but also as a potential oncogene in advanced prostate cancers.

LINC00472 suppresses oral squamous cell carcinoma growth by targeting miR-455-3p/ELF3 axis

ABSTRACT LINC00472 is reported to play a role in suppressing tumors in cancers such as lung cancer and hepatocellular carcinoma, among others. We made investigations into the effects of LINC00472 in oral squamous cell carcinoma (OSCC) progression to explore the underlying molecular mechanisms. By qRT-PCR, we assessed the LINC00472 expression in OSCC tissues and cells and performed functional analysis to investigate how LINC00472/miR-455-3p/ELF3 impacts OSCC cell proliferation, apoptosis, and cell cycle. The role that LINC00472 plays in OSCC tumor growth was examined by establishing a xenograft model. Down-regulation of LINC00472 occurred in tissues and cells of an OSCC tumor. LINC00472 overexpression caused OSCC cell proliferation to be inhibited, cell apoptosis to be promoted, and cell cycle arrest to be induced. As a competing endogenous RNA (ceRNA), LINC00472 can block miR-455-3p function and further promote ELF3 expression. The overexpression of miR-455-3p or ELF3 knockdown was shown to be capable of reversing the anti-tumor effects of LINC00472 in OSCC. In vivo experiments confirmed the tumor-suppressing role of LINC00472 in the progression of OSCC. In short, we found that the novel LINC00472 inhibits OSCC growth via the miR-455-3p/ELF3 axis. LINC00472 and its targeted miR-455-3p/ELF3 axis may represent valuable targets for treating OSCC.

The Immune-Related Gene ELF3 is a Novel Biomarker for the Prognosis of Ovarian Cancer.

BackgroundOvarian cancer (OC) is a fatal gynaecological malignancy. The study aimed to conduct a comprehensive study to determine the role of ELF3 in OC through bioinformatic analysis.MethodsKruskal–Wallis test, Wilcoxon sign-rank test, and logistic regression were used to evaluate the relationship between clinical characteristics and ELF3 expression. Kaplan–Meier method and Cox regression analysis were used to evaluate the prognostic factors. Gene set enrichment analysis (GSEA) and immuno-infiltration analysis were used to evaluate the significant involvement of ELF3 in function.ResultsHigh ELF3 expression in OC was associated with age (P< 0.001). High ELF3 expression predicted a poorer overall survival (OS) (HR: 1.37; 95% CI: 1.05–1.78; P=0.019) and disease specific survival (DSS) (HR: 1.43; 95% CI: 1.08–1.89; P=0.013). And ELF3 expression (HR: 1.779; 95% CI: 1.281–2.472; P<0.001) was independently correlated with OS in OC patients. GSEA demonstrated that pathways including GPCR-ligand binding, neuronal system, signaling by WNT, translation, neuroactive ligand-receptor interaction, and TCF dependent signaling in response to WNT were differentially enriched in ELF3 low expression phenotype. Immune infiltration analysis showed that ELF3 expression was correlated with immune infiltrates.ConclusionELF3 expression in OC patients was significantly associated with poor survival and immune infiltration and a promising prognostic biomarker in OC.

ELF3 activated by a superenhancer and an autoregulatory feedback loop is required for high-level HLA-C expression on extravillous trophoblasts

HLA-C arose during evolution of pregnancy in the great apes 10 to 15 million years ago. It has a dual function on placental extravillous trophoblasts (EVTs) as it contributes to both tolerance and immunity at the maternal-fetal interface. The mode of its regulation is of considerable interest in connection with the biology of pregnancy and pregnancy abnormalities. First-trimester primary EVTs in which HLA-C is highly expressed, as well as JEG3, an EVT model cell line, were employed. Single-cell RNA-seq data and quantitative PCR identified high expression of the transcription factor ELF3 in those cells. Chromatin immunoprecipitation (ChIP)-PCR confirmed that both ELF3 and MED1 bound to the proximal HLA-C promoter region. However, binding of RFX5 to this region was absent or severely reduced, and the adjacent HLA-B locus remained closed. Expression of HLA-C was inhibited by ELF3 small interfering RNAs (siRNAs) and by wrenchnolol treatment. Wrenchnolol is a cell-permeable synthetic organic molecule that mimics ELF3 and is relatively specific for binding to ELF3's coactivator, MED23, as our data also showed in JEG3. Moreover, the ELF3 gene is regulated by a superenhancer that spans more than 5 Mb, identified by assay for transposase-accessible chromatin using sequencing (ATAC-seq), as well as by its sensitivity to (+)-JQ1 (inhibitor of BRD4). ELF3 bound to its own promoter, thus creating an autoregulatory feedback loop that establishes expression of ELF3 and HLA-C in trophoblasts. Wrenchnolol blocked binding of MED23 to ELF3, thus disrupting the positive-feedback loop that drives ELF3 expression, with down-regulation of HLA-C expression as a consequence.

ELF3 Overexpression as Prognostic Biomarker for Recurrence of Stage II Colorectal Cancer.

Adjuvant chemotherapy for high-risk Stage II colorectal cancer (CRC) is weakly recommended; however, no consensus exists on "high-risk" definition. Prognostic biomarker identification is important for selecting patients with poor prognosis who may benefit from adjuvant chemotherapy. Using Microarray data analyses, ELF3 was identified as a candidate gene highly expressed in Stage II CRC with distant recurrences. ELF3 mRNA expression in 168 Stage II CRC patients was subjected to quantitative RT-PCR analysis and ELF3 protein expression in 185 patients was quantified by immunohistochemical analysis. The relationship between mRNA and protein expression levels and patient characteristics were also investigated. The overall recurrence rate and relapse-free survival were significantly poorer in the ELF3 high-expression than the low-expression group at the mRNA and protein levels. High ELF3 mRNA and protein expression levels were independent poor prognostic factors. High ELF3 expression was associated with recurrence of Stage II.

The miR-1224-5p/ELF3 Axis Regulates Malignant Behaviors of Pancreatic Cancer via PI3K/AKT/Notch Signaling Pathways.

PurposeAberrant expression of microRNAs contributes to the progression of pancreatic cancer by targeting downstream genes. A novel regulatory axis, miR-1224-5p/ELF3, was identified by bioinformatic analysis and experimental verification. Studies of the underlying molecular mechanisms behind this axis lead to a better understanding of the development of pancreatic cancer.Materials and MethodsThe differential expression of miR-1224-5p and ELF3 was verified based on Gene Expression Omnibus (GEO) datasets and clinical samples. The relationship between miR-1224-5p and ELF3 was demonstrated by luciferase assay and Western blot. The related signaling pathways of the miR-1224-5p/ELF3 axis in pancreatic cancer were investigated by gene set enrichment analysis (GSEA) and verified by Western blot. An analysis between ELF3 expression and immune infiltration was performed. Cellular and animal experiments were utilized to explore the effects of miR-1224-5p and ELF3 in pancreatic cancer.ResultsSuppressed expression of miR-1224-5p in pancreatic tumor tissues and cancer cells was identified first. Furthermore, miR-1224-5p is correlated with clinicopathological features, and decreased expression of miR-1224-5p indicates poor prognosis. miR-1224-5p serves as a tumor suppressor and inhibits malignant behaviors of pancreatic cancer based on in vivo and in vitro assays. The putative target gene ELF3 was predicted by bioinformatic analysis and confirmed by dual-luciferase reporter assay. Overexpression of ELF3 can improve the malignant behaviors of pancreatic cancer and demonstrates poor prognosis and advanced clinical stage. The inhibitory role of miR-1224-5p in pancreatic cancer is manifested by its direct targeting of ELF3. A negative correlation between ELF3 expression and immune cell infiltration was identified, suggesting an immunosuppressive state resulting from ELF3 overexpression. The PI3K/AKT/Notch signaling pathways and epithelial-to-mesenchymal transition (EMT) are important underlying mechanisms.ConclusionThe miR-1224-5p/ELF3 axis may serve as a new diagnostic, therapeutic, and prognostic biomarker in pancreatic cancer. The related PI3K/AKT/Notch/EMT signaling pathways greatly promote the elucidation of the progression of pancreatic cancer.

Epithelial tumor suppressor ELF3 is a lineage-specific amplified oncogene in lung adenocarcinoma

Gene function in cancer is often cell type-specific. The epithelial cell-specific transcription factor ELF3 is a documented tumor suppressor in many epithelial tumors yet displays oncogenic properties in others. Here, we show that ELF3 is an oncogene in the adenocarcinoma subtype of lung cancer (LUAD), providing genetic, functional, and clinical evidence of subtype specificity. We discover a region of focal amplification at chromosome 1q32.1 encompassing the ELF3 locus in LUAD which is absent in the squamous subtype. Gene dosage and promoter hypomethylation affect the locus in up to 80% of LUAD analyzed. ELF3 expression was required for tumor growth and a pan-cancer expression network analysis supports its subtype and tissue specificity. We further show that ELF3 displays strong prognostic value in LUAD but not LUSC. We conclude that, contrary to many other tumors of epithelial origin, ELF3 is an oncogene and putative therapeutic target in LUAD.

Abstract 5212: Ablation of SMAD4 repression on Elf3 promotes the progression of lung cancer in vivo

Abstract Lung cancer is the leading cause of cancer death among both men and women in United States. ELF3, a transcription factor, overexpresses in human and mouse lung cancer and plays an oncogenic role in lung cancer cells1,2. Understanding the regulatory mechanisms for ELF3 may help to develop related target therapies. Ablation of Smad4 and Pten in mice results in an increase of Elf3 expression and the development of adeno-squamous carcinoma1. Results from in vivo ChIP-Seq analysis demonstrate SMAD4 occupancy near the Elf3 promoter in the pulmonary epithelium1. Therefore, we hypothesize that this SMAD4 occupancy is required to repress Elf3 expression and subsequently suppresses lung cancer progression. To test this hypothesis, we deleted this SMAD4 binding region in vivo and characterized the phenotypic consequences. CRISPR/Cas9 was used to ablate the SMAD4 ChIP-Seq binding peak on Elf3 promoter region in mice. This resulted in mice with 12 and 17 nucleotide deletions in the Elf3 promoter region. ChIP-qPCR analysis showed that SMAD4 binding was decreased while H3K27ac bindings were increased in the Elf3 promoter region. The reduction in SMAD4 binding and the epigenetic changes in the Elf3 promoter region resulted in an increased expression of ELF3 in these lungs of these mice. Histological analysis of these mouse lungs showed the presence of pulmonary hyperplasia and the development of lung tumors. In order to identify potential interacting factors with SMAD4 in the Elf3 promoter, motif analysis of the deleted SMAD4 binding peak identified GATA2, ETS1 and OXT2 as potential interacting factors. In summary, small disruption of Elf3 promoter region bound by SMAD4 leads to the increase of ELF3 expression with the enhanced binding of H3K27ac and the decrease of SMAD4 binding and promotes lung hyperplasia and lung cancer development. SMAD4 may interact with GATA2, ETS2 and OXT2 in the regulation of Elf3expression. Key references 1. Liu, J. et al. ErbB2 Pathway Activation upon Smad4 Loss Promotes Lung Tumor Growth and Metastasis. Cell Rep, doi:10.1016/j.celrep.2015.02.014 (2015). 2. Wang, H. et al. Overexpression of ELF3 facilitates cell growth and metastasis through PI3K/Akt and ERK signaling pathways in non-small cell lung cancer. Int J Biochem Cell Biol 94, 98-106, doi:10.1016/j.biocel.2017.12.002 (2018). Note: This abstract was not presented at the meeting. Citation Format: Jian Liu, Tianyuan Wang, San-pin Wu, Jian-liang Li, Francesco J. Demayo. Ablation of SMAD4 repression on Elf3 promotes the progression of lung cancer in vivo [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 5212.