Ets Family Research Articles

Downregulation of key B cell transcription factor Ets1 in response to antigen receptor signaling depends on IKK2 and unconventional downstream effectors.

Abstract The transcription factor (TF) Ets1 plays a critical role in maintaining B cell quiescence. High Ets1 levels in resting B cells prevent premature plasma cell differentiation and suppress autoimmune responses, but Ets1 must be downregulated to allow for proper B cell responses. Despite this vital role, it remains unknown how the Ets1 expression pattern is coordinated. Upon activation of antigen receptors (ARs), Ets1 is downregulated and quantification of newly transcribed pre-mRNA indicates that a decrease in Ets1 gene transcription is responsible. New protein synthesis is shown to be dispensable for this downregulation, suggesting that it does not rely on upregulation of a repressor but rather changes in abundance or activity of existing TFs. NFκB is activated in response to AR signaling and its binding motifs are found in putative regulatory elements identified near the Ets1 gene. Inhibition of NFκB-activating kinase IKK2 prevents the decrease in Ets1 transcription in response to AR stimulation. However, neither RelA nor cRel downstream of IKK2 are required for proper control of Ets1 expression. Next, we examined potential roles for Foxo3 and Rictor, unconventional downstream effectors of IKK2. Foxo3 has a similar expression pattern to Ets1 and is downregulated upon BCR stimulation, but B cells lacking Foxo3 regulate Ets1 expression in a manner similar to WT B cells. Rictor is a component of the mTORC2 complex, but inhibition of mTOR signaling does not impact Ets1 downregulation in response to BCR stimulation. Overall, these data suggest that Ets1 expression is coordinated by unconventional, but as of yet unidentified, downstream effectors of IKK2 in response to AR signaling. Funded by NIH R01 AI122720 and the Lupus Research Alliance.

Ets1 governs the differential formation of circulating and tissue-resident memory CD8 T cells

Abstract During infections, naive CD8 T cells differentiate into terminal effector cells (TE) and long-lived memory CD8 T cells that recirculate through lymphoid organs using the vasculature (TCIRC), and tissue resident memory (TRM) cells that do not recirculate. The transcriptional control that delineates differentiation of TE cells from memory cell subsets is still unclear. DNA motifs recognized by ETS-family transcription factors (TFs) are highly frequent in cis-regulatory regions that become chromatin accessible in naïve CD8 T cells 24 hours after TCR stimulation, and in mature memory T cell subsets. The ETS-family is encoded by 27 genes, and expression analyses revealed Ets1 is the most highly expressed member. Ets1 is transiently downregulated during activation and is re-expressed in TCIRC cells, but remains lower in intestinal TRM cells. P14 CD8 T cells responding to acute LCMV infection that were depleted of Ets1 using retrovirally-delivered shRNAmirs preferentially became KLRG1hi CD127lo TE-like cells, and their numbers in the circulation rapidly declined, resulting in an absence of long-lived effector cells and residual numbers of central memory-like cells. Concomitantly, Ets1-depleted cells re-distributed into the gut TRM compartment and had increased fractions of CD69hi CD103hi cells. Early after infection in the spleen, a larger fraction of Ets1-depleted P14 cells express CCR9 and α4β7. Depletion or overexpression of Ets1 in P14 cells both indicate it negatively regulates CD25 and Prdm1 expression, and promotes Tcf7 and Bcl6 expression. These results suggest that Ets1 functions early during CD8 T cell activation to establish the normal pattern of TCIRC and TRM cell differentiation during acute viral infection.

SPDEF targets EMT pathway to inhibit cell migration and invasion in triple‐negative breast cancer

Background Current treatments against triple-negative breast cancer (TNBC) are ineffective due to the high rate of metastasis and therapy resistance. There is, therefore, a pressing need for novel and efficacious therapies against TNBC. The basal-like TNBC has a poor prognosis and a high metastasis rate compared to the luminal-like breast cancer subtypes. Here, set to determine the role of Sam Pointed Domain Ets Transcription Factor also known as Prostate-Derived Ets Factor (SPDEF/PDEF), in TNBC. Methods Clinical data were extracted from the METABRIC breast cancer project, hosted at the TCGA database. MDA-MB-231 and MDA 231-LM2-4175 cells were stably transfected with PDEF/control pBABE retroviral vectors. Cell migration/invasion assays were done to explore the effects of PDEF expression on the motility of TNBC cells. Proliferation was measured under 2D and 3D culture conditions. qRT-PCR and immunoblot were performed to visualize gene and protein expression. Data were graphed and analyzed using Graph Pad Prism Software 8 using one-way ANOVA and the unpaired two-tailed Student's t-test. All data were evaluated in triplicate against vector control cells. Results Basal gene and protein expression screening revealed that the expression of PDEF is significantly lower in highly metastatic breast cancer cells compared to the non-metastatic ones. Data analysis from clinical breast cancer METABRIC cohort revealed the loss of PDEF is associated with tumor metastasis, tumor grade, and poor patient survival. PDEF is lost in highly metastatic and therapy resistant TNBC cells. Re-expression of PDEF diminished cell migration and invasion in TNBC cells. Decreased migration and invasion in response to PDEF was not a result of decreased cell growth and proliferation or increased cell death but a result of suppression of EMT markers in TNBC cells. Conclusions PDEF expression inhibits cell migration and invasion partially by down-regulating EMT-related protein markers, suggesting a critical role for PDEF in suppressing TNBC cell metastasis.

ERG transcription factors have a splicing regulatory function involving RBFOX2 that is altered in the EWS-FLI1 oncogenic fusion

ERG family proteins (ERG, FLI1 and FEV) are a subfamily of ETS transcription factors with key roles in physiology and development. In Ewing sarcoma, the oncogenic fusion protein EWS-FLI1 regulates both transcription and alternative splicing of pre-messenger RNAs. However, whether wild-type ERG family proteins might regulate splicing is unknown. Here, we show that wild-type ERG proteins associate with spliceosomal components, are found on nascent RNAs, and induce alternative splicing when recruited onto a reporter minigene. Transcriptomic analysis revealed that ERG and FLI1 regulate large numbers of alternative spliced exons (ASEs) enriched with RBFOX2 motifs and co-regulated by this splicing factor. ERG and FLI1 are associated with RBFOX2 via their conserved carboxy-terminal domain, which is present in EWS-FLI1. Accordingly, EWS-FLI1 is also associated with RBFOX2 and regulates ASEs enriched in RBFOX2 motifs. However, in contrast to wild-type ERG and FLI1, EWS-FLI1 often antagonizes RBFOX2 effects on exon inclusion. In particular, EWS-FLI1 reduces RBFOX2 binding to the ADD3 pre-mRNA, thus increasing its long isoform, which represses the mesenchymal phenotype of Ewing sarcoma cells. Our findings reveal a RBFOX2-mediated splicing regulatory function of wild-type ERG family proteins, that is altered in EWS-FLI1 and contributes to the Ewing sarcoma cell phenotype.

Combination Flavopiridol and SN38 drives toxicity and loss of DNA damage repair in Ewing Sarcoma

EWS-Fli1 is an oncogenic transcription factor that causes Ewing sarcoma. EWS-Fli1 is a fusion protein comprised of an N-terminal low complexity domain from the RNA-binding protein EWSR1 and the C-terminal DNA-binding domain of the ETS transcription factor Fli1. EWS-Fli1 drives expression of oncogenes and decreases expression of tumor suppressor proteins. EWS-Fli1 acts as a transcriptional regulator by recruiting RNA Pol II to promoter regions. Ewing sarcoma is particularly susceptible to DNA damaging chemotherapy compared to other cancers. However, the underlying reason for this sensitivity to DNA damage has not been elucidated. Given the role of EWS-Fli1 in disrupting transcription, we hypothesized that EWS-Fli1 is concurrently disrupting the DNA damage response. During transcription, RNA Pol II is phosphorylated by CDK9 to escape initiation into elongation. We tested the ability of transcription inhibition to affect the DNA damage response by combining treatment a CDK9 inhibitor, flavopiridol, and the DNA damage agent, SN38. This combination drastically diminished cell viability in Ewing cell lines. This sensitivity was not found in non-Ewing cell lines U2OS and HEK293T/17. We also find in Ewing cells that the combination treatment decreases the DNA damage markers gamma-H2A.X and phospho-CHK1 compared to non-Ewing's cells. Together, these results suggest Ewing sarcoma cells are sensitized to DNA damage due in part to transcriptional dysregulation and suggest combination CDK9 inhibition with DNA damage treatment could provide a therapeutic approach for Ewing Sarcoma.

Unique Presentation and Diagnostic Challenges in Metastatic Cerebral Angiosarcoma: Case Report

Abstract BACKGROUND AND IMPORTANCE Angiosarcoma is a malignant endothelial tumor arising from vascular or lymphatic tissue. Metastatic cerebral angiosarcoma has been rarely reported and is associated with a poor prognosis. CLINICAL PRESENTATION We report a 22-yr-old man who presented with a complex partial seizure. Contrasted brain magnetic resonance imaging (MRI) revealed 3 enhancing masses of the left frontal-parietal junction, right occipital lobe, and right caudate nucleus. Computed tomography (CT) of the chest, abdomen, and pelvis and a cardiac MRI revealed no evidence of primary tumor. Immunohistochemistry following biopsy was positive for the ETS transcription factor ERG, CD31, FLI-1, and FosB, confirming angiosarcoma. CT/positron emission tomography (PET) scan showed skeletal metastases in the left iliac bone, sacrum, and T6 vertebral body. The patient underwent resection of the 2 cortical masses, stereotactic radiosurgery to the intracranial lesions as well as to his oligometastases, and was treated with 9 cycles of paclitaxel and bevacizumab. Brain MRI and CT/PET body imaging after cycle 9 revealed no tumor recurrence or progression. The progression-free and overall survival following the diagnosis of brain metastases was 27 mo. CONCLUSION Neurosurgeons should be aware of the unique presentation and diagnostic challenges in metastatic cerebral angiosarcoma, particularly in cases when the primary tumor site is unknown. Furthermore, combined surgery, ablative radiation, and targeted systemic and biological therapy can result in prolonged survival even in patients with metastases.

Reverse Engineering of Ewing Sarcoma Regulatory Network Uncovers PAX7 and RUNX3 as Master Regulators Associated with Good Prognosis.

Simple SummaryEwing Sarcoma is a rare cancer that, when localized, has an overall five-year survival rate of 70%. Patients with metastasis have a five-year survival rate of 15 to 30%. Early analysis of patient prognosis can be crucial to provide adequate treatment and increase chances of survival. Besides, it is a disease with several gaps in our understanding, including regulation of genes and which transcription factors are master regulators. This work addresses these two topics by inferring gene regulatory networks that allow us to identify putative master regulators to predict patient prognosis. We were able to identify two sets of master regulators that can predict good and bad patient outcomes.Ewing Sarcoma (ES) is a rare malignant tumor occurring most frequently in adolescents and young adults. The ES hallmark is a chromosomal translocation between the chromosomes 11 and 22 that results in an aberrant transcription factor (TF) through the fusion of genes from the FET and ETS families, commonly EWSR1 and FLI1. The regulatory mechanisms behind the ES transcriptional alterations remain poorly understood. Here, we reconstruct the ES regulatory network using public available transcriptional data. Seven TFs were identified as potential MRs and clustered into two groups: one composed by PAX7 and RUNX3, and another composed by ARNT2, CREB3L1, GLI3, MEF2C, and PBX3. The MRs within each cluster act as reciprocal agonists regarding the regulation of shared genes, regulon activity, and implications in clinical outcome, while the clusters counteract each other. The regulons of all the seven MRs were differentially methylated. PAX7 and RUNX3 regulon activity were associated with good prognosis while ARNT2, CREB3L1, GLI3, and PBX3 were associated with bad prognosis. PAX7 and RUNX3 appear as highly expressed in ES biopsies and ES cell lines. This work contributes to the understanding of the ES regulome, identifying candidate MRs, analyzing their methilome and pointing to potential prognostic factors.

Decoding molecular markers and transcriptional circuitry of naive and primed states of human pluripotency

Pluripotent stem cells (PSCs) have been observed to occur in two distinct states — naive and primed. Both naive and primed state PSCs can give rise to tissues of all the three germ layers in vitro but differ in their potential to generate germline chimera in vivo. Understanding the molecular mechanisms that govern these two states of pluripotency in human can open a plethora of opportunities for studying early embryonic development and in biomedical applications. In this work, we use weighted gene co-expression network analysis (WGCNA) to identify the key molecular makers and their interactions that define the two distinct pluripotency states. Signed hybrid network was reconstructed from transcriptomic data (RNA-seq) of naive and primed state pluripotent samples. Our analysis revealed two sets of genes that are involved in the establishment and maintenance of naive and primed states. The naive state genes were found to be enriched for biological processes and pathways related to metabolic processes while primed state genes were associated with system development. We further filtered these lists to identify the intra-modular hubs and the hub transcription factors (TFs) for each group. Validation of the identified TFs was carried out using independent microarray datasets and we finally present a list of 52 and 33 TFs as the set of core TFs that are responsible for the induction and maintenance of naive and primed states of pluripotency in human, respectively. Among these, the TFs ZNF275, ZNF232, SP4, and MSANTD3 could be of interest as they were not reported in previous studies.

ER71/ETV2 Promotes Hair Regeneration from Chemotherapeutic Drug-Induced Hair Loss by Enhancing Angiogenesis.

Chemotherapy-induced alopecia and hair loss can be stressful in patients with cancer. The hair grows back, but sometimes the hair tends to stay thin. Therefore, understanding mechanisms regulating hair regeneration may improve the management of chemotherapy-induced alopecia. Previous studies have revealed that chemotherapeutic agents induce a hair follicle vascular injury. As hair growth is associated with micro-vessel regeneration, we postulated that the stimulation of angiogenesis might enhance hair regeneration. In particular, mice treated with 5-fluorouracil (5-FU) showed delayed anagen initiation and reduced capillary density when compared with untreated controls, suggesting that the retardation of anagen initiation by 5-FU treatment may be attributed to the loss of perifollicular micro-vessels. We investigated whether the ETS transcription factor ETV2 (aka ER71), critical for vascular development and regeneration, can promote angiogenesis and hair regrowth in a 5-FU-induced alopecia mouse model. Tie2-Cre; Etv2 conditional knockout (CKO) mice, which lack Etv2 in endothelial cells, presented similar hair regrowth rates as the control mice after depilation. Following 5-FU treatment, Tie2-Cre; Etv2 CKO mice revealed a significant reduction in capillary density, anagen induction, and hair restoration when compared with controls. Mice receiving lentiviral Etv2 injection after 5-FU treatment showed significantly improved anagen induction and hair regrowth. Two-photon laser scanning microscopy revealed that enforced Etv2 expression restored normal vessel morphology after 5-FU mediated vessel injury. Our data suggest that vessel regeneration strategies may improve hair regrowth after chemotherapeutic treatment.

A role for sustained MAPK activity in the mouse ventral telencephalon

The MAPK pathway is a major growth signal that has been implicated during the development of progenitors, neurons, and glia in the embryonic brain. Here, we show that the MAPK pathway plays an important role in the generation of distinct cell types from progenitors in the ventral telencephalon. Our data reveal that phospho-p44/42 (called p-ERK1/2) and the ETS transcription factor Etv5, both downstream effectors in the MAPK pathway, show a regional bias in expression during ventral telencephalic development, with enriched expression in the dorsal region of the LGE and ventral region of the MGE at E13.5 and E15.5. Interestingly, expression of both factors becomes more uniform in ventricular zone (VZ) progenitors by E18.5. To gain insight into the role of MAPK activity during progenitor cell development, we used a cre inducible constitutively active MEK1 allele (RosaMEK1DD/+) in combination with a ventral telencephalon enriched cre (Gsx2e-cre) or a dorsal telencephalon enriched cre (Emx1cre/+). Sustained MEK/MAPK activity in the ventral telencephalon (Gsx2e-cre; RosaMEK1DD/+) expanded dorsal lateral ganglionic eminence (dLGE) enriched genes (Gsx2 and Sp8) and oligodendrocyte progenitor cell (OPC) markers (Olig2, Pdgfrα, and Sox10), and also reduced markers in the ventral (v) LGE domain (Isl1 and Foxp1). Activation of MEK/MAPK activity in the dorsal telencephalon (Emx1cre/+; RosaMEK1DD/+) did not initially activate the expression of dLGE or OPC genes at E15.5 but ectopic expression of Gsx2 and OPC markers were observed at E18.5. These results support the idea that MAPK activity as readout by p-ERK1/2 and Etv5 expression is enriched in distinct subdomains of ventral telencephalic progenitors during development. In addition, sustained activation of the MEK/MAPK pathway in the ventral or dorsal telencephalon influences dLGE and OPC identity from progenitors.

Long noncoding RNA HBBP1 enhances γ-globin expression through the ETS transcription factor ELK1

β-Thalassemia is an autosomal recessive genetic disease caused by defects in the production of adult hemoglobin (HbA, α2β2), which leads to an imbalance between α- and non-α-globin chains. Reactivation of γ-globin expression is an effective strategy to treat β-thalassemia patients. Previously, it was demonstrated that hemoglobin subunit beta pseudogene 1 (HBBP1) is associated with elevated fetal hemoglobin (HbF, α2γ2) in β-thalassemia patients. However, the mechanism underlying HBBP1-mediated HbF production is unknown. In this study, using bioinformatics analysis, we found that HBBP1 is involved in γ-globin production, and then preliminarily confirmed this finding in K562 cells. When HBBP1 was overexpressed, γ-globin expression was increased at the transcript and protein levels in HUDEP-2 cells. Next, we found that ETS transcription factor ELK1 (ELK1) binds to the HBBP1 proximal promoter and significantly promotes its activity. Moreover, the synthesis of γ-globin was enhanced when ELK1 was overexpressed in HUDEP-2 cells. Surprisingly, ELK1 also directly bound to and activated the γ-globin proximal promoter. Furthermore, we found that HBBP1 and ELK1 can interact with each other in HUDEP-2 cells. Collectively, these findings suggest that HBBP1 can induce γ-globin by enhancing ELK1 expression, providing some clues for γ-globin reactivation in β-thalassemia.

Acetylation of ELF5 suppresses breast cancer progression by promoting its degradation and targeting CCND1

E74-like ETS transcription factor 5 (ELF5) is involved in a wide spectrum of biological processes, e.g., mammogenesis and tumor progression. We have identified a list of p300-interacting proteins in human breast cancer cells. Among these, ELF5 was found to interact with p300 via acetylation, and the potential acetylation sites were identified as K130, K134, K143, K197, K228, and K245. Furthermore, an ELF5-specific deacetylase, SIRT6, was also identified. Acetylation of ELF5 promoted its ubiquitination and degradation, but was also essential for its antiproliferative effect against breast cancer, as overexpression of wild-type ELF5 and sustained acetylation-mimicking ELF5 mutant could inhibit the expression of its target gene CCND1. Taken together, the results demonstrated a novel regulation of ELF5 as well as shedding light on its important role in modulation of breast cancer progression.

In vivo CD8+ T cell CRISPR screening reveals control by Fli1 in infection and cancer

Improving effector activity of antigen-specific Tcells is a major goal in cancer immunotherapy. Despite the identification of several effector Tcell (TEFF)-driving transcription factors (TFs), the transcriptional coordination of TEFF biology remains poorly understood. We developed an invivo Tcell CRISPR screening platform and identified a key mechanism restraining TEFF biology through the ETS family TF, Fli1. Genetic deletion of Fli1 enhanced TEFF responses without compromising memory or exhaustion precursors. Fli1 restrained TEFF lineage differentiation by binding to cis-regulatory elements of effector-associated genes. Loss of Fli1 increased chromatin accessibility at ETS:RUNX motifs, allowing more efficient Runx3-driven TEFF biology. CD8+ Tcells lacking Fli1 provided substantially better protection against multiple infections and tumors. These data indicate that Fli1 safeguards the developing CD8+ Tcell transcriptional landscape from excessive ETS:RUNX-driven TEFF cell differentiation. Moreover, genetic deletion of Fli1 improves TEFF differentiation and protective immunity in infections and cancer.

ELK1-induced upregulation lncRNA LINC02381 accelerates the osteosarcoma tumorigenesis through targeting CDCA4 via sponging miR-503–5p

Long noncoding RNAs (lncRNAs) have been identified as functional modulators in human tumors. The purpose of our study was to determine the expressing trend, clinical significance and functions of lncRNA LINC02381(LINC02381) in osteosarcoma. We observed that the expression of LINC02381 and cell division cycle-associated protein 4 (CDCA4) were distinctly increased in osteosarcoma specimens and cells, while miR-503–5p expression was decreased. Additionally, ETS transcription factor ELK1 (ELK1) could bind directly to the LINC02381 promoter region and activate its transcription. Clinical assays revealed that high LINC02381 was associated with advanced clinical progress and poor clinical outcome. Functionally, knockdown of LINC02381 suppressed the proliferation, migration and invasion of osteosarcoma cells. What’s more, LINC02381 could down-regulate CDCA4 via sponging miR-503–5p, and there existed a negative correlation between LINC02381 expression and miR-503–5p expression in 92 osteosarcoma samples. Rescue experiments proved the carcinogenic role of LINC02381/miR-503–5p/CDCA4 axis in osteosarcoma progression. Overall, our data illustrated how LINC02381 played an oncogenic role in osteosarcoma and might offer a novel diagnostic and prognostic biomarker and potential therapeutic target for osteosarcoma.

Gene Regulatory Network of ETS Domain Transcription Factors in Different Stages of Glioma.

The ETS domain family of transcription factors is involved in a number of biological processes, and is commonly misregulated in various forms of cancer. Using microarray datasets from patients with different grades of glioma, we have analyzed the expression profiles of various ETS genes, and have identified ETV1, ELK3, ETV4, ELF4, and ETV6 as novel biomarkers for the identification of different glioma grades. We have further analyzed the gene regulatory networks of ETS transcription factors and compared them to previous microarray studies, where Elk-1-VP16 or PEA3-VP16 were overexpressed in neuroblastoma cell lines, and we identify unique and common regulatory networks for these ETS proteins.

Intestinal Goblet Cell Loss during Chorioamnionitis in Fetal Lambs: Mechanistic Insights and Postnatal Implications.

Chorioamnionitis, an important cause of preterm birth, is linked to necrotizing enterocolitis (NEC). NEC is characterized by a disrupted mucus barrier, goblet cell loss, and endoplasmic reticulum (ER) stress of the intestinal epithelium. These findings prompted us to investigate the mechanisms underlying goblet cell alterations over time in an ovine chorioamnionitis model. Fetal lambs were intra-amniotically (IA) exposed to lipopolysaccharides (LPS) for 5, 12, or 24 h, or 2, 4, 8, or 15 d before premature delivery at 125 d gestational age (GA). Gut inflammation, the number, distribution, and differentiation of goblet cells, ER stress, and apoptosis were measured. We found a biphasic reduction in goblet cell numbers 24 h–2 d after, and 15 d after IA LPS exposure. The second decrease of goblet cell numbers was preceded by intestinal inflammation, apoptosis, and crypt ER stress, and increased SAM-pointed domain-containing ETS transcription factor (SPDEF)-positive cell counts. Our combined findings indicated that ER stress drives apoptosis of maturating goblet cells during chorioamnionitis, ultimately reducing goblet cell numbers. As similar changes have been described in patients suffering from NEC, these findings are considered to be clinically important for understanding the predecessors of NEC, and targeting ER stress in this context is interesting for future therapeutics.

Tet2 Inactivation Enhances the Antitumor Activity of Tumor-Infiltrating Lymphocytes.

Inactivation of tumor-infiltrating lymphocytes (TIL) is one of the mechanisms mitigating antitumor immunity during tumor onset and progression. Epigenetic abnormalities are regarded as a major culprit contributing to the dysfunction of TILs within tumor microenvironments. In this study, we used a murine model of melanoma to discover that Tet2 inactivation significantly enhances the antitumor activity of TILs with an efficacy comparable to immune checkpoint inhibition imposed by anti-PD-L1 treatment. Single-cell RNA-sequencing analysis suggested that Tet2-deficient TILs exhibit effector-like features. Transcriptomic and ATAC-sequencing analysis showed that Tet2 ablation reshapes chromatin accessibility and favors binding of transcription factors geared toward CD8+ T-cell activation. Furthermore, the ETS family of transcription factors contributed to augmented CD8+ T-cell function following Tet2 depletion. Overall, our study establishes that Tet2 constitutes one of the epigenetic barriers that account for dysfunction of TILs and that Tet2 inactivation could promote antitumor immunity to suppress tumor growth. SIGNIFICANCE: This study suggests that ablation of TET2+ from TILs could promote their antitumor function by reshaping chromatin accessibility for key transcription factors and enhancing the transcription of genes essential for antitumor activity.

Higher ETV5 Expression Associates With Poor 5-Florouracil-Based Adjuvant Therapy Response in Colon Cancer.

Discovery of markers predictive for 5-Fluorouracil (5-FU)-based adjuvant chemotherapy (adjCTX) response in patients with locally advanced stage II and III colon cancer (CC) is necessary for precise identification of potential therapy responders. PEA3 subfamily of ETS transcription factors (ETV1, ETV4, and ETV5) are upregulated in multiple cancers including colon cancers. However, the underlying epigenetic mechanism regulating their overexpression as well as their role in predicting therapy response in colon cancer are largely unexplored. In this study, using gene expression and methylation data from The Cancer Genome Atlas (TCGA) project, we showed that promoter DNA methylation negatively correlates with ETV4 expression (ρ = −0.17, p = 5.6 × 10–3) and positively correlates with ETV5 expression (ρ = 0.22, p = 1.43 × 10–4) in colon cancer tissue. Further, our analysis in 1,482 colon cancer patients from five different cohorts revealed that higher ETV5 expression associates with shorter relapse-free survival (RFS) of adjCTX treated colon cancer patients (Hazard ratio = 2.09–5.43, p = 0.004–0.01). The present study suggests ETV5 expression as a strong predictive biomarker for 5-FU-based adjCTX response in stage II/III CC patients.

Regulation of peripheral Th/Treg differentiation and suppression of airway inflammation by Nr4a transcription factors

SummaryHelper T (Th) and regulatory T (Treg) cell differentiation programs promote the eradication of pathogens, while minimizing adverse immune reactions. Here, we found that Nr4a family of nuclear receptors supports Treg cell induction and represses Th1 and Th2 cell differentiation from naive CD4+ T cells. Nr4a factors are transiently induced in CD4+ T cells immediately after antigen stimulation, thereby mediating epigenetic changes. In differentiating Treg cells, Nr4a factors mainly upregulated the early responsive genes in the Treg cell-specifying gene set, either directly or in cooperation with Ets family transcription factors. In contrast, Nr4a factors repressed AP-1 activity by interrupting a positive feedback loop for Batf factor expression, thus suppressing Th2 cell-associated genes. In an allergic airway inflammation model, Nr4a factors suppressed the pathogenesis, mediating oral tolerance. Lastly, pharmacological activation of an engineered Nr4a molecule prevented allergic airway inflammation, indicating that Nr4a factors may be novel therapeutic targets for inflammatory diseases.

Pluripotent stem cell-derived epithelium misidentified as brain microvascular endothelium requires ETS factors to acquire vascular fate

Cells derived from pluripotent sources in vitro must resemble those found in vivo as closely as possible at both transcriptional and functional levels in order to be a useful tool for studying diseases and developing therapeutics. Recently, differentiation of human pluripotent stem cells (hPSCs) into brain microvascular endothelial cells (ECs) with blood-brain barrier (BBB)-like properties has been reported. These cells have since been used as a robust in vitro BBB model for drug delivery and mechanistic understanding of neurological diseases. However, the precise cellular identity of these induced brain microvascular endothelial cells (iBMECs) has not been well described. Employing a comprehensive transcriptomic metaanalysis of previously published hPSC-derived cells validated by physiological assays, we demonstrate that iBMECs lack functional attributes of ECs since they are deficient in vascular lineage genes while expressing clusters of genes related to the neuroectodermal epithelial lineage (Epi-iBMEC). Overexpression of key endothelial ETS transcription factors (ETV2, ERG, and FLI1) reprograms Epi-iBMECs into authentic endothelial cells that are congruent with bona fide endothelium at both transcriptomic as well as some functional levels. This approach could eventually be used to develop a robust human BBB model in vitro that resembles the human brain EC in vivo for functional studies and drug discovery.