Pioneer Transcription Factor Research Articles

MECHANISMS IN ENDOCRINOLOGY: Pioneer transcription factors in pituitary development and tumorigenesis.

Pioneer transcription factors have key roles in development as master regulators of cell fate specification. Only a small fraction of all transcription factors have the pioneer ability that confers access to target genomic DNA sites embedded in so-called 'closed' heterochromatin. This ability to seek and bind target sites within the silenced portion of the epigenome is the basis for their role in changing cell fate. Upon binding heterochromatin sites, pioneer factors trigger remodeling of chromatin from a repressed into an active organization. This action is typically exerted at enhancer regulatory sequences, thus allowing activation of new gene subsets. During pituitary development, the only pioneer with a well-documented role is Pax7 that specifies the intermediate lobe melanotrope cell fate. In this review, a particular focus is placed on this Pax7 function but its properties are also considered within the general context of pioneer factor action. Given their potent activity to reprogram gene expression, it is not surprising that many pioneers are associated with tumor development. Overexpression or chromosomal translocations leading to the production of chimeric pioneers have been implicated in different cancers. We review here the current knowledge on the mechanism of pioneer factor action.

PRDM1 Drives Human Primary T Cell Hyporesponsiveness by Altering the T Cell Transcriptome and Epigenome

Background: T cell hyporesponsiveness is crucial for the functional immune system and prevents the damage induced by alloreactive T cells in autoimmune pathology and transplantation. Methods: We detected the phenotype of PRDM1 overexpressed human primary T cells. To investigate the molecular mechanisms of PRDM1 regulating T cells, we performed RNA-seq, MethylationEPIC BeadChip array, CUT&Tag and ATAC-seq in PRDM1 overexpressed human primary T cells. Findings: Overexpression of PRDM1 in primary T cells expanded Treg cell subsets and increased the secretion of IL-4, while decreased expression had the opposite effects. Integrated multiomics analysis showed that PRDM1 directly upregulated T cell inhibitory genes such as CD244, KLF2 and KLRD1 and downregulated the T cell activation gene IL2, indicating that PRDM1 could promote a tolerant transcriptional profile in primary T cells. Binding motifs of key T cell function regulators, such as FOS, JUN and AP-1, were enriched in PRDM1 binding sites and that PRDM1 altered the chromatin accessibility of these regions. Furthermore, the PRDM1 expression level in allografts was negatively related to acute graft-versus-host disease (aGVHD) occurrence after hematopoietic stem cell transplantation (HSCT). Interpretation: Our systematic multiomics analysis found that the transcription factor (TF) PRDM1 acts as a pioneer TF and master regulator during T cell hyporesponsiveness in human primary T cells. These results demonstrated that PRDM1 is sufficient for inducing T cell hyporesponsiveness and could be a predictive biomarker for HSCT prognosis. Funding: This work was partly supported by grants from the National Key Research and Development Program of China (2017YFA0104500), Innovative Research Groups of the National Natural Science Foundation of China (No. 81621001), Key Program of the National Natural Science Foundation of China (No.81530046, 81930004), Beijing Municipal Natural Science Foundation (Grant No. 7204319). Declaration of Interest: None to declare. Ethical Approval: The study has been approved by the Ethics Committee of Peking University People's Hospital, and written informed consent from all subjects was obtained

Emerging role of pioneer transcription factors in targeted ERα positive breast cancer.

Transcription factors (TFs) are modular protein groups that preferably bind to DNA sequences and guide genomic expression through transcription. Among these key regulators, “pioneer factors” are an emerging class of TFs that specifically interact with nucleosomal DNA and facilitate accessible genomic binding sites for the additional TFs. There is growing evidence of these specialized modulators in particular malignancies, as highlighted by agents’ clinical efficacy, specifically targeting nuclear hormone receptors. They have been implicated in multiple cancers more recently, with a high proportion inculpating on hormone influential cancers. Moreover, extended crosstalk and cooperation between ERα pioneering factors in estrogen-dependent breast cancer (BC) remain elucidated. This review discusses on the recent advances in our understanding of pioneer TFs in cancer, especially highlighting its potentiality to modulate chromatin condensation to permit ERα recruitment in BC cells. Through the study it was concluded that the highly prospected pioneer TFs in BC, including FOXA1, TLE1, PBX1, and GATA3, possess the potential therapeutic significance and further innovations in the field could yield targeted therapy in cancer treatment.

WAPL maintains a cohesin loading cycle to preserve cell-type-specific distal gene regulation.

The cohesin complex plays an essential role in maintaining genome organization. However, its role in gene regulation remains largely unresolved. Here, we report that the cohesin release factor WAPL creates a pool of free cohesin, in a process known as cohesin turnover, which reloads it to cell-type specific binding sites. Paradoxically, stabilization of cohesin binding, following WAPL ablation, results in depletion of cohesin from these cell-type specific regions, a loss of gene expression and differentiation. Chromosome conformation capture experiments show that cohesin turnover is important for maintaining promoter-enhancer loops. Binding of cohesin to cell-type specific sites is dependent on the pioneer transcription factors OCT4 (POU5F1) and SOX2, but not NANOG. We show the importance of cohesin turnover in controlling transcription and propose that a cycle of cohesin loading and off-loading, instead of static cohesin binding, mediates promoter and enhancer interactions critical for gene regulation.

Pioneer-like factor GAF cooperates with PBAP (SWI/SNF) and NURF (ISWI) to regulate transcription.

Transcriptionally silent genes must be activated throughout development. This requires nucleosomes be removed from promoters and enhancers to allow transcription factor (TF) binding and recruitment of coactivators and RNA polymerase II (Pol II). Specialized pioneer TFs bind nucleosome-wrapped DNA to perform this chromatin opening by mechanisms that remain incompletely understood. Here, we show that GAGA factor (GAF), a Drosophila pioneer-like factor, functions with both SWI/SNF and ISWI family chromatin remodelers to allow recruitment of Pol II and entry to a promoter-proximal paused state, and also to promote Pol II's transition to productive elongation. We found that GAF interacts with PBAP (SWI/SNF) to open chromatin and allow Pol II to be recruited. Importantly, this activity is not dependent on NURF as previously proposed; however, GAF also synergizes with NURF downstream from this process to ensure efficient Pol II pause release and transition to productive elongation, apparently through its role in precisely positioning the +1 nucleosome. These results demonstrate how a single sequence-specific pioneer TF can synergize with remodelers to activate sets of genes. Furthermore, this behavior of remodelers is consistent with findings in yeast and mice, and likely represents general, conserved mechanisms found throughout eukarya.

It’s a Phase That EBF1 Is Going Through

EBF1 is a pioneer transcription factor involved in B lymphocyte specification. In this issue of Immunity, Wang etal. localize EBF1's pioneering activity to a prion-like domain that mediates recruitment of the nucleosome remodeler Brg1 and FUS-assisted liquid-liquid phase separation.

Relationships between Breast Feeding and Breast Cancer Subtypes: Lessons Learned from Studies in Humans and in Mice.

There are differential risk relationships between parity and breast cancer according to estrogen receptor (ER) status, with an increased risk of ER- disease reduced by breastfeeding. This may be particularly relevant for understanding the higher incidence of ER- tumors in Black women, who are more likely to be parous and less likely to breastfeed than other U.S. groups. Potential mechanisms for these relationships may include effects of disordered breast involution on inflammatory milieu in the breast as well as epigenetic reprogramming in the mammary gland, which can affect cell fate decisions in progenitor cell pools. In normal breast tissue, parity has been associated with hypermethylation of FOXA1, a pioneer transcription factor that promotes the luminal phenotype in luminal progenitors, while repressing the basal phenotype. In breast tumors, relationships between FOXA1 methylation and parity were strongest among women who did not breastfeed. Here, we summarize the epidemiologic literature regarding parity, breastfeeding, and breast cancer subtypes, and review potential mechanisms whereby these factors may influence breast carcinogenesis, with a focus on effects on progenitor cell pools in the mammary gland.

A sensitive mNeonGreen reporter system to measure transcriptional dynamics in Drosophila development

The gene regulatory network governing anterior–posterior axis formation in Drosophila is a well-established paradigm to study transcription in developmental biology. The rapid temporal dynamics of gene expression during early stages of development, however, are difficult to track with standard techniques. We optimized the bright and fast-maturing fluorescent protein mNeonGreen as a real-time, quantitative reporter of enhancer expression. We derive enhancer activity from the reporter fluorescence dynamics with high spatial and temporal resolution, using a robust reconstruction algorithm. By comparing our results with data obtained with the established MS2-MCP system, we demonstrate the higher detection sensitivity of our reporter. We used the reporter to quantify the activity of variants of a simple synthetic enhancer, and observe increased activity upon reduction of enhancer–promoter distance or addition of binding sites for the pioneer transcription factor Zelda. Our reporter system constitutes a powerful tool to study spatio-temporal gene expression dynamics in live embryos.

Abstract PO-099: SOX2 delineates a mouse lung adenocarcinoma subtype vulnerable to targeted therapy

Abstract Lung adenocarcinomas comprise the largest fraction of non-small cell lung cancer, which is the leading cause of cancer deaths. 75% of adenocarcinomas lack targeted therapies due to scarcity of druggable drivers. We leveraged transcriptional data from >800 early-stage and advanced patients to classify tumors based on signaling similarities and discover subgroups within this unmet patient population. The subtypes capture heterogeneity even amongst tumors lacking known oncogenic drivers. Paired multi-regional intratumoral biopsies demonstrate unified subtypes despite divergently evolved pro-oncogenic mutations, indicating subtype stability during selective pressure. We identified differential subtype response to MEK inhibition across multiple preclinical model systems and a clinical trial, supporting prognostic utility of transcriptional subtyping. Differential subtype dependency on MEK signaling reproduced in a mouse model of KRAS-mutant lung adenocarcinoma, where a MEK-dependent adenocarcinoma subtype is driven by a SOX2 cellular state. Our findings support forward translational relevance of transcriptional subtypes and reveal that naturally evolved yet ectopic expression of a pioneer transcription factor may modulate tumor subtype and response. Citation Format: Jonathan E. Cooper, Anneleen Daemen, Dorothee Nickles, Szymon Myrta, Oded Foreman, Jeff Eastham, Melissa R. Junttila, Heinrich Jasper. SOX2 delineates a mouse lung adenocarcinoma subtype vulnerable to targeted therapy [abstract]. In: Proceedings of the AACR Virtual Special Conference on Tumor Heterogeneity: From Single Cells to Clinical Impact; 2020 Sep 17-18. Philadelphia (PA): AACR; Cancer Res 2020;80(21 Suppl):Abstract nr PO-099.

Structural Basis of Inhibition of the Pioneer Transcription Factor NF-Y by Suramin.

NF-Y is a transcription factor (TF) comprising three subunits (NF-YA, NF-YB, NF-YC) that binds with high specificity to the CCAAT sequence, a widespread regulatory element in gene promoters of prosurvival, cell-cycle-promoting, and metabolic genes. Tumor cells undergo “metabolic rewiring” through overexpression of genes involved in such pathways, many of which are under NF-Y control. In addition, NF-YA appears to be overexpressed in many tumor types. Thus, limiting NF-Y activity may represent a desirable anti-cancer strategy, which is an ongoing field of research. With virtual-screening docking simulations on a library of pharmacologically active compounds, we identified suramin as a potential NF-Y inhibitor. We focused on suramin given its high water-solubility that is an important factor for in vitro testing, since NF-Y is sensitive to DMSO. By electrophoretic mobility shift assays (EMSA), isothermal titration calorimetry (ITC), STD NMR, X-ray crystallography, and molecular dynamics (MD) simulations, we showed that suramin binds to the histone fold domains (HFDs) of NF-Y, preventing DNA-binding. Our analyses, provide atomic-level detail on the interaction between suramin and NF-Y and reveal a region of the protein, nearby the suramin-binding site and poorly conserved in other HFD-containing TFs, that may represent a promising starting point for rational design of more specific and potent inhibitors with potential therapeutic applications.

A small protein encoded by a putative lncRNA regulates apoptosis and tumorigenicity in human colorectal cancer cells.

Long noncoding RNAs (lncRNAs) are often associated with polysomes, indicating coding potential. However, only a handful of endogenous proteins encoded by putative lncRNAs have been identified and assigned a function. Here, we report the discovery of a putative gastrointestinal-tract-specific lncRNA (LINC00675) that is regulated by the pioneer transcription factor FOXA1 and encodes a conserved small protein of 79 amino acids which we termed FORCP (FOXA1-Regulated Conserved Small Protein). FORCP transcript is undetectable in most cell types but is abundant in well-differentiated colorectal cancer (CRC) cells where it functions to inhibit proliferation, clonogenicity, and tumorigenesis. The epitope-tagged and endogenous FORCP protein predominantly localizes to the endoplasmic reticulum (ER). In response to ER stress, FORCP depletion results in decreased apoptosis. Our findings on the initial characterization of FORCP demonstrate that FORCP is a novel, conserved small protein encoded by a mis-annotated lncRNA that regulates apoptosis and tumorigenicity in well-differentiated CRC cells.

The Transcription Factor TCF1 in T Cell Differentiation and Aging.

The transcription factor T cell factor 1 (TCF1), a pioneer transcription factor as well as a downstream effector of WNT/β-catenin signaling, is indispensable for T cell development in the thymus. Recent studies have highlighted the additional critical role of TCF1 in peripheral T cell responses to acute and chronic infections as well as cancer. Here, we review the regulatory functions of TCF1 in the differentiation of T follicular helper cells, memory T cells and recently described stem-like exhausted T cells, where TCF1 promotes less differentiated stem-like cell states by controlling common gene-regulatory networks. These studies also provide insights into the mechanisms of defective T cell responses in older individuals. We discuss alterations in TCF1 expression and related regulatory networks with age and their consequences for T cell responses to infections and vaccination. The increasing understanding of the pathways regulating TCF1 expression and function in aged T cells holds the promise of enabling the design of therapeutic interventions aiming at improving T cell responses in older individuals.

Pioneer Transcription Factors Initiating Gene Network Changes.

Pioneer transcription factors have the intrinsic biochemical ability to scan partial DNA sequence motifs that are exposed on the surface of a nucleosome and thus access silent genes that are inaccessible to other transcription factors. Pioneer factors subsequently enable other transcription factors, nucleosome remodeling complexes, and histone modifiers to engage chromatin, thereby initiating the formation of an activating or repressive regulatory sequence. Thus, pioneer factors endow the competence for fate changes in embryonic development, are essential for cellular reprogramming, and rewire gene networks in cancer cells. Recent studies with reconstituted nucleosomes in vitro and chromatin binding in vivo reveal that pioneer factors can directly perturb nucleosome structure and chromatin accessibility in different ways. This review focuses on our current understanding of the mechanisms by which pioneer factors initiate gene network changes and will ultimately contribute to our ability to control cell fates at will.

FOXA1 Mutations Reveal Distinct Chromatin Profiles and Influence Therapeutic Response in Breast Cancer

Mutations in the pioneer transcription factor FOXA1 are a hallmark of estrogen receptor-positive (ER+) breast cancers. Examining FOXA1 in ∼5,000 breast cancer patients identifies several hotspot mutations in the Wing2 region and a breast cancer-specific mutation SY242CS, located in the third β strand. Using a clinico-genomically curated cohort, together with breast cancer models, we find that FOXA1 mutations associate with a lower response to aromatase inhibitors. Mechanistically, Wing2 mutations display increased chromatin binding at ER loci upon estrogen stimulation, and an enhanced ER-mediated transcription without changes in chromatin accessibility. In contrast, SY242CS shows neomorphic properties that include the ability to open distinct chromatin regions and activate an alternative cistrome and transcriptome. Structural modeling predicts that SY242CS confers a conformational change that mediates stable binding to a non-canonical DNA motif. Taken together, our results provide insights into how FOXA1 mutations perturb its function to dictate cancer progression and therapeutic response.

Chromatin binding of FOXA1 is promoted by LSD1-mediated demethylation in prostate cancer.

FOXA1 functions as a pioneer transcription factor by facilitating the access to chromatin for steroid hormone receptors, such as androgen receptor and estrogen receptor1-4, but mechanisms regulating its binding to chromatin remain elusive. LSD1 (KDM1A) acts as a transcriptional repressor by demethylating mono/dimethylated histone H3 lysine 4 (H3K4me1/2)5,6, but also acts as a steroid hormone receptor coactivator through mechanisms that are unclear. Here we show, in prostate cancer cells, that LSD1 associates with FOXA1 and active enhancer markers, and that LSD1 inhibition globally disrupts FOXA1 chromatin binding. Mechanistically, we demonstrate that LSD1 positively regulates FOXA1 binding by demethylating lysine 270, adjacent to the wing2 region of the FOXA1 DNA-binding domain. Acting through FOXA1, LSD1 inhibition broadly disrupted androgen-receptor binding and its transcriptional output, and dramatically decreased prostate cancer growth alone and in synergy with androgen-receptor antagonist treatment in vivo. These mechanistic insights suggest new therapeutic strategies in steroid-driven cancers.

Interaction of the pioneer transcription factor GATA3 with nucleosomes

During cellular reprogramming, the pioneer transcription factor GATA3 binds chromatin, and in a context-dependent manner directs local chromatin remodeling and enhancer formation. Here, we use high-resolution nucleosome mapping in human cells to explore the impact of the position of GATA motifs on the surface of nucleosomes on productive enhancer formation, finding productivity correlates with binding sites located near the nucleosomal dyad axis. Biochemical experiments with model nucleosomes demonstrate sufficiently stable transcription factor-nucleosome interaction to empower cryo-electron microscopy structure determination of the complex at 3.15 Å resolution. The GATA3 zinc fingers efficiently bind their target 5′-GAT-3′ sequences in the nucleosome when they are located in solvent accessible, consecutive major grooves without significant changes in nucleosome structure. Analysis of genomic loci bound by GATA3 during reprogramming suggests a correlation of recognition motif sequence and spacing that may distinguish productivity of new enhancer formation.

Alternative Activation of Macrophages Is Accompanied by Chromatin Remodeling Associated with Lineage-Dependent DNA Shape Features Flanking PU.1 Motifs.

IL-4 activates macrophages to adopt distinct phenotypes associated with clearance of helminth infections and tissue repair, but the phenotype depends on the cellular lineage of these macrophages. The molecular basis of chromatin remodeling in response to IL-4 stimulation in tissue-resident and monocyte-derived macrophages is not understood. In this study, we find that IL-4 activation of different lineages of peritoneal macrophages in mice is accompanied by lineage-specific chromatin remodeling in regions enriched with binding motifs of the pioneer transcription factor PU.1. PU.1 motif is similarly associated with both tissue-resident and monocyte-derived IL-4-induced accessible regions but has different lineage-specific DNA shape features and predicted cofactors. Mutation studies based on natural genetic variation between C57BL/6 and BALB/c mouse strains indicate that accessibility of these IL-4-induced regions can be regulated through differences in DNA shape without direct disruption of PU.1 motifs. We propose a model whereby DNA shape features of stimulation-dependent genomic elements contribute to differences in the accessible chromatin landscape of alternatively activated macrophages on different genetic backgrounds that may contribute to phenotypic variations in immune responses.

Abstract 3437: Cistromically-dominant class2 mutants of FOXA1 de-repress WNT signaling to drive prostate cancer metastasis

Abstract Forkhead box A1 (FOXA1) is a pioneer transcription factor that is essential for the normal development of several endoderm-derived organs, including the prostate gland. FOXA1 independently binds to and de-compacts condensed chromatin to reveal the binding sites of partnering nuclear hormone receptors. In prostate luminal epithelial cells, FOXA1 delimits tissue-specific enhancers that are transcriptionally activated by the androgen receptor (AR). Thus, it is intriguing that FOXA1 is frequently mutated in hormone-receptor-driven prostate, breast, bladder and salivary-gland tumors. In a recent study, leveraging an aggregate cohort of 1,546 prostate cancers, we classified FOXA1 alterations into three novel structural classes that diverge in clinical incidence and genetic co-alteration profiles, with a collective prevalence of 35%. Class1 mutations comprise of missense and in-frame indels within the Wing2 secondary structure and originate in early prostate cancer without other driver alterations in genes such as ETS or SPOP. Contrarily, class2 mutations comprise of frameshift mutations which truncate the C-terminal end of the protein and are clonally acquired in the metastatic disease. Finally, class-3 genomic rearrangements comprise of duplications and translocations that reposition a conserved enhancer element to drive overexpression of FOXA1 or other oncogenes. Here, we mechanistically expand upon the neomorphic class2 mutants and show that they centrally enable prostate cancer metastasis. Remarkably, we found truncated class2 mutants to entirely displace their wild-type counterpart from the chromatin - a characteristic we termed as “cistromic dominance” - due to higher DNA affinity. Through proteomic assays, we identified the C-terminal regulatory domain of wild-type FOXA1 to interact with and recruit a bonafide transcriptional WNT-repressor called TLE3 to the chromatin. Notably, class2 mutants had lost this interaction and dominantly displace TLE3 from chromatin to de-repress WNT signaling. Accordingly, we found CRISPR-engineered prostate cancer cells harboring class2 FOXA1 mutations to show higher invasive and metastatic ability both in vitro and in vivo. Also, the increase in the metastatic ability of class2-mutant cells was completely reversed upon treatment with a WNT inhibitor. Overall, our study reaffirms the central role of FOXA1 in mediating AR-driven oncogenesis and provides mechanistic insights into how distinct classes of FOXA1 alterations drive initiation or metastatic progression of prostate cancer. These results reveal a novel FOXA1-TLE3-WNT signaling axis and rationalize its therapeutic co-inhibition in advanced prostate cancer. Citation Format: Abhijit Parolia, Marcin Cieslik, Sandra Carson, Shih-Chun Chu, Lanbo Xiao, Takahiro Ouchi, Mustapha Jaber, Yuping Zhang, Pankaj Vats, Xuhong Cao, Arul Chinnaiyan. Cistromically-dominant class2 mutants of FOXA1 de-repress WNT signaling to drive prostate cancer metastasis [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 3437.

Abstract 1305: Structure-function mapping reveal FLI1 contributes more than the DNA-binding properties to drive Ewing sarcoma biology

Abstract Objective: Ewing sarcoma (ES) is the second most common pediatric bone cancer. To address a blatant lack of advancement in treatment, it is of the utmost importance to understand the underlying biology driving disease and through these developments, we hope to better outcomes through bench-to-bedside projects. ES is driven by a chromosomal translocation, which produces a fusion oncoprotein known as EWS/FLI1. Though it is known that FLI1 contains a DNA-binding domain that is essential for protein function and disease formation, very little further research has been completed to understand precisely how it contributes to ES etiology and how we can disrupt these functions. Our goal is to complete structure-function mapping that allows an in-depth analysis of FLI1 domain contributions that ultimately may elicit new targets for treatment development. Methods: EWS/FLI1 cDNA was used to create several different versions of EWS/FLI1, including a full-length protein and two EWS/FLI1 proteins with truncated FLI1 domains. A ES cellular model was used to test functionality of these constructs. A variety of molecular biology and new sequencing techniques were employed to establish structure-function relationships of the FLI1 domain of EWS/FLI1, including luciferase reporter assays, soft agar assays, RNA-sequencing, CUT&RUN-sequencing, and ATAC-sequencing. Results: In vitro transcriptional activation assays revealed EWS/FLI1 containing only the DNA-binding domain of FLI1 (EWS/FLI1DBD) was sufficient to drive transcription, but this result did not translate to the ability to oncogenically transform ES cells in a soft agar assay. We used various sequencing techniques to understand the process that allows EWS/FLI1 to actually drive transformation. CUT&RUN-sequencing revealed that EWS/FLI1 binds various places throughout the genome and, surprisingly, that EWS/FLI1DBD was able to bind all the same places. RNA-sequencing showed that EWS/FLI1 was able to regulate gene expression at the majority of bound regions, but EWS/FLI1DBD starkly lacked this ability. These results caused us to question what role EWS/FLI1 has as a pioneer transcription factor and what happens between DNA-binding and gene regulation. Through ATAC-sequencing results, we expect to see that EWS/FLI1 has the ability to open closed chromatin regions, but that EWS/FLI1DBD will lack this ability. It is likely that EWS/FLI1 recruits various additional co-factors to open chromatin and through these constructs, we hope to identify more of these important proteins. Conclusions: We have revealed FLI1 has a novel role in EWS/FLI1 activity that goes beyond DNA-binding, as well as isolated the essential regions of FLI1 necessary for activity. Ultimately, we hope that these novel properties or interactions will provide new ways to target ES cells that can benefit patient outcomes. Citation Format: Megann A. Boone, Jesse C. Crow, Julia Selich-Anderson, Cenny Taslim, Emily R. Theisen, Stephen L. Lessnick. Structure-function mapping reveal FLI1 contributes more than the DNA-binding properties to drive Ewing sarcoma biology [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 1305.

Pioneer transcription factors, progesterone receptor and Krüppel like transcription factor 4, cooperatively stimulate the bovine herpesvirus 1 ICP0 early promoter and productive late protein expression

Bovine herpesvirus 1 (BoHV-1), including commercially available modified live vaccines, readily infect the fetus and ovaries, which can cause reproductive failure. The BoHV-1 latency-reactivation cycle in sensory neurons further complicates reproductive failure because progesterone sporadically induces reactivation from latency. The progesterone receptor (PR) and Krüppel-like transcription factor 15 (KLF15) cooperatively stimulate productive infection and the immediate early transcription unit 1 (IEtu1) promoter. In addition to the IEtu1 promoter, the bICP0 gene also contains a separate early (E) promoter. In this study, we tested the hypothesis that PR and KLF family members transactivate the bICP0 E promoter. PR and KLF4 stimulated bICP0 E promoter activity and expression of late productive viral protein expression in a cooperative manner. Additional studies revealed three enhancer domains within the bICP0 E promoter were responsive to PR and KLF4. Chromatin immunoprecipitation studies demonstrated PR and KLF4 occupy bICP0 E promoter sequences in transfected Neuro-2A cells and at late times following infection of bovine kidney cells. Co-immunoprecipitation studies indicated PR and KLF4 stably interact with each other. These studies suggest cooperative activation of the bICP0 E promoter by PR and KLF4 correlate with interactions between these pioneer transcription factors.