Mathieu Lupien | University of Toronto (original) (raw)
Papers by Mathieu Lupien
Proceedings of the National Academy of Sciences of the United States of America, 2013
The estrogen receptor (ER)α drives growth in two-thirds of all breast cancers. Several targeted t... more The estrogen receptor (ER)α drives growth in two-thirds of all breast cancers. Several targeted therapies, collectively termed endocrine therapy, impinge on estrogen-induced ERα activation to block tumor growth. However, half of ERα-positive breast cancers are tolerant or acquire resistance to endocrine therapy. We demonstrate that genome-wide reprogramming of the chromatin landscape, defined by epigenomic maps for regulatory elements or transcriptional activation and chromatin openness, underlies resistance to endocrine therapy. This annotation reveals endocrine therapy-response specific regulatory networks where NOTCH pathway is overactivated in resistant breast cancer cells, whereas classical ERα signaling is epigenetically disengaged. Blocking NOTCH signaling abrogates growth of resistant breast cancer cells. Its activation state in primary breast tumors is a prognostic factor of resistance in endocrine treated patients. Overall, our work demonstrates that chromatin landscape reprogramming underlies changes in regulatory networks driving endocrine therapy resistance in breast cancer.
Epigenomics, 2012
Over two-thirds of breast cancers rely on estrogen receptor α (ERα) for their growth. Endocrine t... more Over two-thirds of breast cancers rely on estrogen receptor α (ERα) for their growth. Endocrine therapies antagonize estrogen-dependent ERα activation but resistance to these treatments occurs and is associated with poor prognosis. Crosstalk between alternative survival pathways and ERα are currently held as the primary cause of resistance. However, blocking these pathways does not cure endocrine therapy resistant breast cancer suggesting the existence of additional mechanisms. While cancer is commonly considered a genetic disease, the importance of epigenetic events in promoting tumor initiation and progression is increasingly recognized. Here, we consider how epigenetic modifications and alterations to the chromatin landscape contribute to endocrine therapy resistance by modulating ERα expression or altering its genomic activity.
Médecine sciences : M/S, 2012
Nature genetics, 2012
Genome-wide association studies (GWAS) have identified thousands of SNPs that are associated with... more Genome-wide association studies (GWAS) have identified thousands of SNPs that are associated with human traits and diseases. But, because the vast majority of these SNPs are located in non-coding regions of the genome, the mechanisms by which they promote disease risk have remained elusive. Employing a new methodology that combines cistromics, epigenomics and genotype imputation, we annotate the non-coding regions of the genome in breast cancer cells and systematically identify the functional nature of SNPs associated with breast cancer risk. Our results show that breast cancer risk-associated SNPs are enriched in the cistromes of FOXA1 and ESR1 and the epigenome of histone H3 lysine 4 monomethylation (H3K4me1) in a cancer- and cell type-specific manner. Furthermore, the majority of the risk-associated SNPs modulate the affinity of chromatin for FOXA1 at distal regulatory elements, thereby resulting in allele-specific gene expression, which is exemplified by the effect of the rs4784227 SNP on the TOX3 gene within the 16q12.1 risk locus.
Journal of the National Cancer Institute, 2012
To identify downstream effectors of MAPK signaling that could be used as potential additional the... more To identify downstream effectors of MAPK signaling that could be used as potential additional therapeutic targets for BRAF V600E inhibitors, we used hTERT/CDK4R24C/p53DDimmortalized primary human melanocytes genetically modified to ectopically express BRAFV600E or NRASG12D and observed induction of the AP-1 transcription factor family member c-Jun. Using a dominant negative approach, in vitro cell proliferation assays, western blots, and flow cytometry showed that MAPK signaling via BRAF V600E promotes melanoma cell proliferation at G1 through AP-1-mediated negative regulation of the INK4 family member, cyclin-dependent kinase inhibitor 2C (CDKN2C), and the CIP/KIP family member, cyclin-dependent kinase inhibitor 1A (CDKN1A). These effects were antagonized by pharmacological inhibition of CDKN2C and CDKN1A targets CDK2 and CDK4 in vitro. In contrast to BRAFV600E or NRASG12D-expressing melanocytes, melanoma cells have an inherent resistance to suppression of AP-1 activity by BRAF V600E -or MEK-inhibitors. Here, CDK2/4 inhibition statistically significantly augmented the effects of BRAF V600E -or MEK-inhibitors on melanoma cell viability in vitro and growth in athymic nude Foxn1 nu mice (P = .03 when mean tumor volume at day 13 was compared for BRAF V600E inhibitor vs BRAF V600E inhibitor plus CDK2/4 inhibition; P = .02 when mean tumor volume was compared for MEK inhibitor vs MEK inhibitor plus CDK2/4 inhibition; P values were calculated by a two-sided Welch t test; n = 4-8 mice per group).
Trends in genetics : TIG, 2012
Genetic association studies have become standard approaches to characterize the genetic and epige... more Genetic association studies have become standard approaches to characterize the genetic and epigenetic variability associated with cancer development, including predispositions and mutations. However, the bewildering genetic and phenotypic heterogeneity inherent in cancer both magnifies the conceptual and methodological problems associated with these approaches and renders difficult the translation of available genetic information into a knowledge that is both biologically sound and clinically relevant. Here, we elaborate on the underlying causes of this complexity, illustrate why it represents a challenge for genetic association studies, and briefly discuss how it can be reconciled with the ultimate goals of identifying targetable disease pathways and successfully treating individual patients.
Genome research, 2012
Genome-wide association studies (GWAS) are identifying genetic predisposition to various diseases... more Genome-wide association studies (GWAS) are identifying genetic predisposition to various diseases. The 17q24.3 locus harbors the single nucleotide polymorphism (SNP) rs1859962 that is statistically associated with prostate cancer (PCa). It defines a 130-kb linkage disequilibrium (LD) block that lies in an~2-Mb gene desert area. The functional biology driving the risk associated with this LD block is unknown. Here, we integrate genome-wide chromatin landscape data sets, namely, epigenomes and chromatin openness from diverse cell types. This identifies a PCa-specific enhancer within the rs1859962 risk LD block that establishes a 1-Mb chromatin loop with the SOX9 gene. The rs8072254 and rs1859961 SNPs mapping to this enhancer impose allele-specific gene expression. The variant allele of rs8072254 facilitates androgen receptor (AR) binding driving increased enhancer activity. The variant allele of rs1859961 decreases FOXA1 binding while increasing AP-1 binding. The latter is key to imposing allele-specific gene expression. The rs8072254 variant in strong LD with the rs1859962 risk SNP can account for the risk associated with this locus, while rs1859961 is a rare variant less likely to contribute to the risk associated with this LD block. Together, our results demonstrate that multiple genetic variants mapping to a unique enhancer looping to the SOX9 oncogene can account for the risk associated with the PCa 17q24.3 locus. Allele-specific recruitment of the transcription factors androgen receptor (AR) and activating protein-1 (AP-1) account for the increased enhancer activity ascribed to this PCa-risk LD block. This further supports the notion that an integrative genomics approach can identify the functional biology disrupted by genetic risk variants.
PloS one, 2012
PBX1 is a TALE homeodomain transcription factor involved in organogenesis and tumorigenesis. Alth... more PBX1 is a TALE homeodomain transcription factor involved in organogenesis and tumorigenesis. Although it has been shown that ovarian, breast, and melanoma cancer cells depend on PBX1 for cell growth and survival, the molecular mechanism of how PBX1 promotes tumorigenesis remains unclear. Here, we applied an integrated approach by overlapping PBX1 ChIP-chip targets with the PBX1-regulated transcriptome in ovarian cancer cells to identify genes whose transcription was directly regulated by PBX1. We further determined if PBX1 target genes identified in ovarian cancer cells were co-overexpressed with PBX1 in carcinoma tissues. By analyzing TCGA gene expression microarray datasets from ovarian serous carcinomas, we found co-upregulation of PBX1 and a significant number of its direct target genes. Among the PBX1 target genes, a homeodomain protein MEOX1 whose DNA binding motif was enriched in PBX1immunoprecipicated DNA sequences was selected for functional analysis. We demonstrated that MEOX1 protein interacts with PBX1 protein and inhibition of MEOX1 yields a similar growth inhibitory phenotype as PBX1 suppression. Furthermore, ectopically expressed MEOX1 functionally rescued the PBX1-withdrawn effect, suggesting MEOX1 mediates the cellular growth signal of PBX1. These results demonstrate that MEOX1 is a critical target gene and cofactor of PBX1 in ovarian cancers.
Science (New York, N.Y.), 2012
PLoS genetics, 2011
Altered transcriptional programs are a hallmark of diseases, yet how these are established is sti... more Altered transcriptional programs are a hallmark of diseases, yet how these are established is still ill-defined. PBX1 is a TALE homeodomain protein involved in the development of different types of cancers. The estrogen receptor alpha (ERα) is central to the development of two-thirds of all breast cancers. Here we demonstrate that PBX1 acts as a pioneer factor and is essential for the ERα-mediated transcriptional response driving aggressive tumors in breast cancer. Indeed, PBX1 expression correlates with ERα in primary breast tumors, and breast cancer cells depleted of PBX1 no longer proliferate following estrogen stimulation. Profiling PBX1 recruitment and chromatin accessibility across the genome of breast cancer cells through ChIP-seq and FAIRE-seq reveals that PBX1 is loaded and promotes chromatin openness at specific genomic locations through its capacity to read specific epigenetic signatures. Accordingly, PBX1 guides ERα recruitment to a specific subset of sites. Expression profiling studies demonstrate that PBX1 controls over 70% of the estrogen response. More importantly, the PBX1-dependent transcriptional program is associated with poor-outcome in breast cancer patients. Correspondingly, PBX1 expression alone can discriminate a priori the outcome in ERα-positive breast cancer patients. These features are markedly different from the previously characterized ERα-associated pioneer factor FoxA1. Indeed, PBX1 is the only pioneer factor identified to date that discriminates outcome such as metastasis in ERα-positive breast cancer patients. Together our results reveal that PBX1 is a novel pioneer factor defining aggressive ERα-positive breast tumors, as it guides ERα genomic activity to unique genomic regions promoting a transcriptional program favorable to breast cancer progression.
Cancer research, 2011
Nuclear receptors (NRs) comprise a superfamily of ligand-activated transcription factors that pla... more Nuclear receptors (NRs) comprise a superfamily of ligand-activated transcription factors that play important roles in both physiology and diseases including cancer. The technologies of Chromatin ImmunoPrecipitation followed by array hybridization (ChIP-chip) or massively parallel sequencing (ChIP-seq) has been used to map, at an unprecedented rate, the in vivo genome-wide binding (cistrome) of NRs in both normal and cancer cells. We developed a curated database of 88 NR cistrome datasets and other associated high-throughput datasets, including 121 collaborating factor cistromes, 94 epigenomes and 319 transcriptomes. Through integrative analysis of the curated NR ChIP-chip/seq datasets, we discovered novel factor-specific noncanonical motifs that may have important regulatory roles. We also revealed a common feature of NR pioneering factors to recognize relatively short and AT-rich motifs. Most NRs bind predominantly to introns and distal intergenetic regions, and binding sites closer to transcription start sites (TSSs) were found to be neither stronger nor more evolutionarily conserved. Interestingly, while most NRs appear to be predominantly transcriptional activators, our analysis suggests that the binding of ESR1, RARA and RARG has both activating and repressive effects. Through meta-analysis of different omic data of the same cancer cell line model from multiple studies, we generated consensus cistrome and expression profiles. We further made probabilistic predictions of the NR target genes by integrating cistrome and transcriptome data, and validated the predictions using expression data from tumor samples. The final database, with comprehensive cistrome, epigenome, transcriptome datasets, and downstream analysis results, constitutes a valuable resource for the nuclear receptor and cancer community.
Trends in genetics : TIG, 2011
Chromatin is a well-known obstacle to transcription as it controls DNA accessibility, which direc... more Chromatin is a well-known obstacle to transcription as it controls DNA accessibility, which directly impacts the recruitment of the transcriptional machinery. The recent burst of functional genomic studies provides new clues as to how transcriptional competency is regulated in this context. In this review, we discuss how these studies have shed light on a specialized subset of transcription factors, defined as pioneer factors, which direct recruitment of downstream transcription factors to establish lineage-specific transcriptional programs. In particular, we present evidence of an interplay between pioneer factors and the epigenome that could be central to this process. Finally, we discuss how pioneer factors, whose expression and function are altered in tumors, are also being considered for their prognostic value and should therefore be regarded as potential therapeutic targets. Thus, pioneer factors emerge as key players that connect the epigenome and transcription in health and disease.
Wiley interdisciplinary reviews. Systems biology and medicine
The conceptual foundation of the genome-wide association study (GWAS) has advanced unchecked sinc... more The conceptual foundation of the genome-wide association study (GWAS) has advanced unchecked since its conception. A revision might seem premature as the potential of GWAS has not been fully realized. Multiple technical and practical limitations need to be overcome before GWAS can be fairly criticized. But with the completion of hundreds of studies and a deeper understanding of the genetic architecture of disease, warnings are being raised. The results compiled to date indicate that risk-associated variants lie predominantly in non-coding regions of the genome. Additionally, alternative methodologies are uncovering large and heterogeneous sets of rare variants underlying disease. The fear is that, even in its fulfilment, the current GWAS paradigm might be incapable of dissecting all kinds of phenotypes. In the following text we review several initiatives that aim to overcome these limitations. The overarching theme of these studies is the inclusion of biological knowledge to both the analysis and interpretation of genotyping data. GWAS is uninformed of biology by design and although there is some virtue in its simplicity it is also its most conspicuous deficiency. We propose a framework in which to integrate these novel approaches, both empirical and theoretical, in the form of a genome-wide regulatory network (GWRN). By processing experimental data into networks, emerging data types based on chromatin-immunoprecipitation are made computationally tractable. This will give GWAS re-analysis efforts the most current and relevant substrates, and root them firmly on our knowledge of human disease.
Genes & development, 2010
Estrogen receptor a (ERa) expression in breast cancer is predictive of response to endocrine ther... more Estrogen receptor a (ERa) expression in breast cancer is predictive of response to endocrine therapy; however, resistance is common in ERa-positive tumors that overexpress the growth factor receptor ERBB2. Even in the absence of estrogen, ERa can be activated by growth factors, including the epidermal growth factor (EGF). EGF induces a transcriptional program distinct from estrogen; however, the mechanism of the stimulus-specific response is unknown. Here we show that the EGF-induced ERa genomic targets, its cistromes, are distinct from those induced by estrogen in a process dependent on the transcription factor AP-1. The EGF-induced ERa cistrome specifically regulates genes found overexpressed in ERBB2-positive human breast cancers. This provides a potential molecular explanation for the endocrine therapy resistance seen in ERa-positive breast cancers that overexpress ERBB2. These results suggest a central role for ERa in hormone-refractory breast tumors dependent on growth factor pathway activation and favors the development of therapeutic strategies completely antagonizing ERa, as opposed to blocking its estrogen responsiveness alone.
Nature genetics, 2010
Chromatin plays a central role in eukaryotic gene regulation. We have performed genome-wide mappi... more Chromatin plays a central role in eukaryotic gene regulation. We have performed genome-wide mapping of epigenetically-marked nucleosomes to determine their position both near transcription start sites and at distal regulatory elements including enhancers. In prostate cancer cells where androgen receptor (AR) binds primarily to enhancers, we found that androgen treatment dismisses a central nucleosome present over AR binding sites that is flanked by a pair of marked nucleosomes. A novel quantitative model built on the behavior of such nucleosome pairs correctly identified regions bound by the regulators of the immediate androgen response including AR and FoxA1. More importantly this model also correctly predicted novel binding sites for other transcription factors present following prolonged androgen stimulation including Oct1 and NKX3.1. Thus quantitative modeling of enhancer structure provides a powerful predictive method to infer the identity of transcription factors involved in cellular responses to specific stimuli.
Proceedings of the National Academy of Sciences of the United States of America, 2010
Gene expression signatures are used in the clinic as prognostic tools to determine the risk of in... more Gene expression signatures are used in the clinic as prognostic tools to determine the risk of individual patients with localized breast tumors developing distant metastasis. We lack a clear understanding, however, of whether these correlative biomarkers link to a common biological network that regulates metastasis. We find that the c-MYC oncoprotein coordinately regulates the expression of 13 different "poor-outcome" cancer signatures. In addition, functional inactivation of MYC in human breast cancer cells specifically inhibits distant metastasis in vivo and invasive behavior in vitro of these cells. These results suggest that MYC oncogene activity (as marked by "poor-prognosis" signature expression) may be necessary for the translocation of poor-outcome human breast tumors to distant sites.
Molecular and cellular biology, 2010
An analysis of mRNA expression in T47D breast cancer cells treated with the synthetic progestin R... more An analysis of mRNA expression in T47D breast cancer cells treated with the synthetic progestin R5020 revealed a subset of progesterone receptor (PR) target genes that are enriched for E2F binding sites. Following up on this observation, we determined that PR-B acts in both direct and indirect manners to positively upregulate E2F1 expression in T47D cells. The direct effects of PR on E2F1 expression were confirmed by chromatin immunoprecipitation (ChIP) analysis, which indicated that the agonist-bound receptor was recruited to several enhancer elements proximal to the E2F1 transcript. However, we also noted that cycloheximide partially inhibits R5020 induction of E2F1 expression, indicating that the ligand-dependent actions of PR on this gene may involve additional indirect regulatory pathways. In support of this hypothesis, we demonstrated that treatment with R5020 significantly increases both hyperphosphorylation of Rb and recruitment of E2F1 to its own promoter, thus activating a positive feedback loop that further amplifies its transcription. Furthermore, we established that PR-mediated induction of Krüppel-like factor 15 (KLF15), which can bind to GC-rich DNA within the E2F1 promoter, is required for maximal induction of E2F1 expression by progestins. Taken together, these results suggest a new paradigm for multimodal regulation of target gene expression by PR.
Proceedings of the National Academy of Sciences of the United States of America, 2013
The estrogen receptor (ER)α drives growth in two-thirds of all breast cancers. Several targeted t... more The estrogen receptor (ER)α drives growth in two-thirds of all breast cancers. Several targeted therapies, collectively termed endocrine therapy, impinge on estrogen-induced ERα activation to block tumor growth. However, half of ERα-positive breast cancers are tolerant or acquire resistance to endocrine therapy. We demonstrate that genome-wide reprogramming of the chromatin landscape, defined by epigenomic maps for regulatory elements or transcriptional activation and chromatin openness, underlies resistance to endocrine therapy. This annotation reveals endocrine therapy-response specific regulatory networks where NOTCH pathway is overactivated in resistant breast cancer cells, whereas classical ERα signaling is epigenetically disengaged. Blocking NOTCH signaling abrogates growth of resistant breast cancer cells. Its activation state in primary breast tumors is a prognostic factor of resistance in endocrine treated patients. Overall, our work demonstrates that chromatin landscape reprogramming underlies changes in regulatory networks driving endocrine therapy resistance in breast cancer.
Epigenomics, 2012
Over two-thirds of breast cancers rely on estrogen receptor α (ERα) for their growth. Endocrine t... more Over two-thirds of breast cancers rely on estrogen receptor α (ERα) for their growth. Endocrine therapies antagonize estrogen-dependent ERα activation but resistance to these treatments occurs and is associated with poor prognosis. Crosstalk between alternative survival pathways and ERα are currently held as the primary cause of resistance. However, blocking these pathways does not cure endocrine therapy resistant breast cancer suggesting the existence of additional mechanisms. While cancer is commonly considered a genetic disease, the importance of epigenetic events in promoting tumor initiation and progression is increasingly recognized. Here, we consider how epigenetic modifications and alterations to the chromatin landscape contribute to endocrine therapy resistance by modulating ERα expression or altering its genomic activity.
Médecine sciences : M/S, 2012
Nature genetics, 2012
Genome-wide association studies (GWAS) have identified thousands of SNPs that are associated with... more Genome-wide association studies (GWAS) have identified thousands of SNPs that are associated with human traits and diseases. But, because the vast majority of these SNPs are located in non-coding regions of the genome, the mechanisms by which they promote disease risk have remained elusive. Employing a new methodology that combines cistromics, epigenomics and genotype imputation, we annotate the non-coding regions of the genome in breast cancer cells and systematically identify the functional nature of SNPs associated with breast cancer risk. Our results show that breast cancer risk-associated SNPs are enriched in the cistromes of FOXA1 and ESR1 and the epigenome of histone H3 lysine 4 monomethylation (H3K4me1) in a cancer- and cell type-specific manner. Furthermore, the majority of the risk-associated SNPs modulate the affinity of chromatin for FOXA1 at distal regulatory elements, thereby resulting in allele-specific gene expression, which is exemplified by the effect of the rs4784227 SNP on the TOX3 gene within the 16q12.1 risk locus.
Journal of the National Cancer Institute, 2012
To identify downstream effectors of MAPK signaling that could be used as potential additional the... more To identify downstream effectors of MAPK signaling that could be used as potential additional therapeutic targets for BRAF V600E inhibitors, we used hTERT/CDK4R24C/p53DDimmortalized primary human melanocytes genetically modified to ectopically express BRAFV600E or NRASG12D and observed induction of the AP-1 transcription factor family member c-Jun. Using a dominant negative approach, in vitro cell proliferation assays, western blots, and flow cytometry showed that MAPK signaling via BRAF V600E promotes melanoma cell proliferation at G1 through AP-1-mediated negative regulation of the INK4 family member, cyclin-dependent kinase inhibitor 2C (CDKN2C), and the CIP/KIP family member, cyclin-dependent kinase inhibitor 1A (CDKN1A). These effects were antagonized by pharmacological inhibition of CDKN2C and CDKN1A targets CDK2 and CDK4 in vitro. In contrast to BRAFV600E or NRASG12D-expressing melanocytes, melanoma cells have an inherent resistance to suppression of AP-1 activity by BRAF V600E -or MEK-inhibitors. Here, CDK2/4 inhibition statistically significantly augmented the effects of BRAF V600E -or MEK-inhibitors on melanoma cell viability in vitro and growth in athymic nude Foxn1 nu mice (P = .03 when mean tumor volume at day 13 was compared for BRAF V600E inhibitor vs BRAF V600E inhibitor plus CDK2/4 inhibition; P = .02 when mean tumor volume was compared for MEK inhibitor vs MEK inhibitor plus CDK2/4 inhibition; P values were calculated by a two-sided Welch t test; n = 4-8 mice per group).
Trends in genetics : TIG, 2012
Genetic association studies have become standard approaches to characterize the genetic and epige... more Genetic association studies have become standard approaches to characterize the genetic and epigenetic variability associated with cancer development, including predispositions and mutations. However, the bewildering genetic and phenotypic heterogeneity inherent in cancer both magnifies the conceptual and methodological problems associated with these approaches and renders difficult the translation of available genetic information into a knowledge that is both biologically sound and clinically relevant. Here, we elaborate on the underlying causes of this complexity, illustrate why it represents a challenge for genetic association studies, and briefly discuss how it can be reconciled with the ultimate goals of identifying targetable disease pathways and successfully treating individual patients.
Genome research, 2012
Genome-wide association studies (GWAS) are identifying genetic predisposition to various diseases... more Genome-wide association studies (GWAS) are identifying genetic predisposition to various diseases. The 17q24.3 locus harbors the single nucleotide polymorphism (SNP) rs1859962 that is statistically associated with prostate cancer (PCa). It defines a 130-kb linkage disequilibrium (LD) block that lies in an~2-Mb gene desert area. The functional biology driving the risk associated with this LD block is unknown. Here, we integrate genome-wide chromatin landscape data sets, namely, epigenomes and chromatin openness from diverse cell types. This identifies a PCa-specific enhancer within the rs1859962 risk LD block that establishes a 1-Mb chromatin loop with the SOX9 gene. The rs8072254 and rs1859961 SNPs mapping to this enhancer impose allele-specific gene expression. The variant allele of rs8072254 facilitates androgen receptor (AR) binding driving increased enhancer activity. The variant allele of rs1859961 decreases FOXA1 binding while increasing AP-1 binding. The latter is key to imposing allele-specific gene expression. The rs8072254 variant in strong LD with the rs1859962 risk SNP can account for the risk associated with this locus, while rs1859961 is a rare variant less likely to contribute to the risk associated with this LD block. Together, our results demonstrate that multiple genetic variants mapping to a unique enhancer looping to the SOX9 oncogene can account for the risk associated with the PCa 17q24.3 locus. Allele-specific recruitment of the transcription factors androgen receptor (AR) and activating protein-1 (AP-1) account for the increased enhancer activity ascribed to this PCa-risk LD block. This further supports the notion that an integrative genomics approach can identify the functional biology disrupted by genetic risk variants.
PloS one, 2012
PBX1 is a TALE homeodomain transcription factor involved in organogenesis and tumorigenesis. Alth... more PBX1 is a TALE homeodomain transcription factor involved in organogenesis and tumorigenesis. Although it has been shown that ovarian, breast, and melanoma cancer cells depend on PBX1 for cell growth and survival, the molecular mechanism of how PBX1 promotes tumorigenesis remains unclear. Here, we applied an integrated approach by overlapping PBX1 ChIP-chip targets with the PBX1-regulated transcriptome in ovarian cancer cells to identify genes whose transcription was directly regulated by PBX1. We further determined if PBX1 target genes identified in ovarian cancer cells were co-overexpressed with PBX1 in carcinoma tissues. By analyzing TCGA gene expression microarray datasets from ovarian serous carcinomas, we found co-upregulation of PBX1 and a significant number of its direct target genes. Among the PBX1 target genes, a homeodomain protein MEOX1 whose DNA binding motif was enriched in PBX1immunoprecipicated DNA sequences was selected for functional analysis. We demonstrated that MEOX1 protein interacts with PBX1 protein and inhibition of MEOX1 yields a similar growth inhibitory phenotype as PBX1 suppression. Furthermore, ectopically expressed MEOX1 functionally rescued the PBX1-withdrawn effect, suggesting MEOX1 mediates the cellular growth signal of PBX1. These results demonstrate that MEOX1 is a critical target gene and cofactor of PBX1 in ovarian cancers.
Science (New York, N.Y.), 2012
PLoS genetics, 2011
Altered transcriptional programs are a hallmark of diseases, yet how these are established is sti... more Altered transcriptional programs are a hallmark of diseases, yet how these are established is still ill-defined. PBX1 is a TALE homeodomain protein involved in the development of different types of cancers. The estrogen receptor alpha (ERα) is central to the development of two-thirds of all breast cancers. Here we demonstrate that PBX1 acts as a pioneer factor and is essential for the ERα-mediated transcriptional response driving aggressive tumors in breast cancer. Indeed, PBX1 expression correlates with ERα in primary breast tumors, and breast cancer cells depleted of PBX1 no longer proliferate following estrogen stimulation. Profiling PBX1 recruitment and chromatin accessibility across the genome of breast cancer cells through ChIP-seq and FAIRE-seq reveals that PBX1 is loaded and promotes chromatin openness at specific genomic locations through its capacity to read specific epigenetic signatures. Accordingly, PBX1 guides ERα recruitment to a specific subset of sites. Expression profiling studies demonstrate that PBX1 controls over 70% of the estrogen response. More importantly, the PBX1-dependent transcriptional program is associated with poor-outcome in breast cancer patients. Correspondingly, PBX1 expression alone can discriminate a priori the outcome in ERα-positive breast cancer patients. These features are markedly different from the previously characterized ERα-associated pioneer factor FoxA1. Indeed, PBX1 is the only pioneer factor identified to date that discriminates outcome such as metastasis in ERα-positive breast cancer patients. Together our results reveal that PBX1 is a novel pioneer factor defining aggressive ERα-positive breast tumors, as it guides ERα genomic activity to unique genomic regions promoting a transcriptional program favorable to breast cancer progression.
Cancer research, 2011
Nuclear receptors (NRs) comprise a superfamily of ligand-activated transcription factors that pla... more Nuclear receptors (NRs) comprise a superfamily of ligand-activated transcription factors that play important roles in both physiology and diseases including cancer. The technologies of Chromatin ImmunoPrecipitation followed by array hybridization (ChIP-chip) or massively parallel sequencing (ChIP-seq) has been used to map, at an unprecedented rate, the in vivo genome-wide binding (cistrome) of NRs in both normal and cancer cells. We developed a curated database of 88 NR cistrome datasets and other associated high-throughput datasets, including 121 collaborating factor cistromes, 94 epigenomes and 319 transcriptomes. Through integrative analysis of the curated NR ChIP-chip/seq datasets, we discovered novel factor-specific noncanonical motifs that may have important regulatory roles. We also revealed a common feature of NR pioneering factors to recognize relatively short and AT-rich motifs. Most NRs bind predominantly to introns and distal intergenetic regions, and binding sites closer to transcription start sites (TSSs) were found to be neither stronger nor more evolutionarily conserved. Interestingly, while most NRs appear to be predominantly transcriptional activators, our analysis suggests that the binding of ESR1, RARA and RARG has both activating and repressive effects. Through meta-analysis of different omic data of the same cancer cell line model from multiple studies, we generated consensus cistrome and expression profiles. We further made probabilistic predictions of the NR target genes by integrating cistrome and transcriptome data, and validated the predictions using expression data from tumor samples. The final database, with comprehensive cistrome, epigenome, transcriptome datasets, and downstream analysis results, constitutes a valuable resource for the nuclear receptor and cancer community.
Trends in genetics : TIG, 2011
Chromatin is a well-known obstacle to transcription as it controls DNA accessibility, which direc... more Chromatin is a well-known obstacle to transcription as it controls DNA accessibility, which directly impacts the recruitment of the transcriptional machinery. The recent burst of functional genomic studies provides new clues as to how transcriptional competency is regulated in this context. In this review, we discuss how these studies have shed light on a specialized subset of transcription factors, defined as pioneer factors, which direct recruitment of downstream transcription factors to establish lineage-specific transcriptional programs. In particular, we present evidence of an interplay between pioneer factors and the epigenome that could be central to this process. Finally, we discuss how pioneer factors, whose expression and function are altered in tumors, are also being considered for their prognostic value and should therefore be regarded as potential therapeutic targets. Thus, pioneer factors emerge as key players that connect the epigenome and transcription in health and disease.
Wiley interdisciplinary reviews. Systems biology and medicine
The conceptual foundation of the genome-wide association study (GWAS) has advanced unchecked sinc... more The conceptual foundation of the genome-wide association study (GWAS) has advanced unchecked since its conception. A revision might seem premature as the potential of GWAS has not been fully realized. Multiple technical and practical limitations need to be overcome before GWAS can be fairly criticized. But with the completion of hundreds of studies and a deeper understanding of the genetic architecture of disease, warnings are being raised. The results compiled to date indicate that risk-associated variants lie predominantly in non-coding regions of the genome. Additionally, alternative methodologies are uncovering large and heterogeneous sets of rare variants underlying disease. The fear is that, even in its fulfilment, the current GWAS paradigm might be incapable of dissecting all kinds of phenotypes. In the following text we review several initiatives that aim to overcome these limitations. The overarching theme of these studies is the inclusion of biological knowledge to both the analysis and interpretation of genotyping data. GWAS is uninformed of biology by design and although there is some virtue in its simplicity it is also its most conspicuous deficiency. We propose a framework in which to integrate these novel approaches, both empirical and theoretical, in the form of a genome-wide regulatory network (GWRN). By processing experimental data into networks, emerging data types based on chromatin-immunoprecipitation are made computationally tractable. This will give GWAS re-analysis efforts the most current and relevant substrates, and root them firmly on our knowledge of human disease.
Genes & development, 2010
Estrogen receptor a (ERa) expression in breast cancer is predictive of response to endocrine ther... more Estrogen receptor a (ERa) expression in breast cancer is predictive of response to endocrine therapy; however, resistance is common in ERa-positive tumors that overexpress the growth factor receptor ERBB2. Even in the absence of estrogen, ERa can be activated by growth factors, including the epidermal growth factor (EGF). EGF induces a transcriptional program distinct from estrogen; however, the mechanism of the stimulus-specific response is unknown. Here we show that the EGF-induced ERa genomic targets, its cistromes, are distinct from those induced by estrogen in a process dependent on the transcription factor AP-1. The EGF-induced ERa cistrome specifically regulates genes found overexpressed in ERBB2-positive human breast cancers. This provides a potential molecular explanation for the endocrine therapy resistance seen in ERa-positive breast cancers that overexpress ERBB2. These results suggest a central role for ERa in hormone-refractory breast tumors dependent on growth factor pathway activation and favors the development of therapeutic strategies completely antagonizing ERa, as opposed to blocking its estrogen responsiveness alone.
Nature genetics, 2010
Chromatin plays a central role in eukaryotic gene regulation. We have performed genome-wide mappi... more Chromatin plays a central role in eukaryotic gene regulation. We have performed genome-wide mapping of epigenetically-marked nucleosomes to determine their position both near transcription start sites and at distal regulatory elements including enhancers. In prostate cancer cells where androgen receptor (AR) binds primarily to enhancers, we found that androgen treatment dismisses a central nucleosome present over AR binding sites that is flanked by a pair of marked nucleosomes. A novel quantitative model built on the behavior of such nucleosome pairs correctly identified regions bound by the regulators of the immediate androgen response including AR and FoxA1. More importantly this model also correctly predicted novel binding sites for other transcription factors present following prolonged androgen stimulation including Oct1 and NKX3.1. Thus quantitative modeling of enhancer structure provides a powerful predictive method to infer the identity of transcription factors involved in cellular responses to specific stimuli.
Proceedings of the National Academy of Sciences of the United States of America, 2010
Gene expression signatures are used in the clinic as prognostic tools to determine the risk of in... more Gene expression signatures are used in the clinic as prognostic tools to determine the risk of individual patients with localized breast tumors developing distant metastasis. We lack a clear understanding, however, of whether these correlative biomarkers link to a common biological network that regulates metastasis. We find that the c-MYC oncoprotein coordinately regulates the expression of 13 different "poor-outcome" cancer signatures. In addition, functional inactivation of MYC in human breast cancer cells specifically inhibits distant metastasis in vivo and invasive behavior in vitro of these cells. These results suggest that MYC oncogene activity (as marked by "poor-prognosis" signature expression) may be necessary for the translocation of poor-outcome human breast tumors to distant sites.
Molecular and cellular biology, 2010
An analysis of mRNA expression in T47D breast cancer cells treated with the synthetic progestin R... more An analysis of mRNA expression in T47D breast cancer cells treated with the synthetic progestin R5020 revealed a subset of progesterone receptor (PR) target genes that are enriched for E2F binding sites. Following up on this observation, we determined that PR-B acts in both direct and indirect manners to positively upregulate E2F1 expression in T47D cells. The direct effects of PR on E2F1 expression were confirmed by chromatin immunoprecipitation (ChIP) analysis, which indicated that the agonist-bound receptor was recruited to several enhancer elements proximal to the E2F1 transcript. However, we also noted that cycloheximide partially inhibits R5020 induction of E2F1 expression, indicating that the ligand-dependent actions of PR on this gene may involve additional indirect regulatory pathways. In support of this hypothesis, we demonstrated that treatment with R5020 significantly increases both hyperphosphorylation of Rb and recruitment of E2F1 to its own promoter, thus activating a positive feedback loop that further amplifies its transcription. Furthermore, we established that PR-mediated induction of Krüppel-like factor 15 (KLF15), which can bind to GC-rich DNA within the E2F1 promoter, is required for maximal induction of E2F1 expression by progestins. Taken together, these results suggest a new paradigm for multimodal regulation of target gene expression by PR.