Marian Waterman - Profile on Academia.edu (original) (raw)

Papers by Marian Waterman

Proceedings of the National Academy of Sciences of the United States of America, Apr 30, 2002

Life science alliance, Jul 13, 2022

Crosstalk between the Hedgehog and MAPK signaling pathways occurs in several types of cancer and ... more Crosstalk between the Hedgehog and MAPK signaling pathways occurs in several types of cancer and contributes to clinical resistance to Hedgehog pathway inhibitors. Here we show that MAP kinase-mediated phosphorylation weakens the binding of the GLI1 transcription factor to its negative regulator SUFU. ERK2 phosphorylates GLI1 on three evolutionarily conserved target sites (S102, S116, and S130) located near the high-affinity binding site for SUFU; these phosphorylations cooperate to weaken the affinity of GLI1-SUFU binding by over 25-fold. Phosphorylation of any one, or even any two, of the three sites does not result in the level of SUFU release seen when all three sites are phosphorylated. Tumor-derived mutations in R100 and S105, residues bordering S102, also diminish SUFU binding, collectively defining a novel evolutionarily conserved SUFU affinity-modulating region. In cultured mammalian cells, GLI1 variants containing phosphomimetic substitutions of S102, S116, and S130 displayed an increased ability to drive transcription. We conclude that multisite phosphorylation of GLI1 by ERK2 or other MAP kinases weakens GLI1-SUFU binding, thereby facilitating GLI1 activation and contributing to both physiological and pathological crosstalk.

Trends in Microbiology, 2008

Nature Immunology, Aug 2, 2009

The differentiation of activated CD4 + T cells into the T helper type 1 (T H 1) or T H 2 fate is ... more The differentiation of activated CD4 + T cells into the T helper type 1 (T H 1) or T H 2 fate is regulated by cytokines and the transcription factors T-bet and GATA-3. Whereas interleukin 12 (IL-12) produced by antigen-presenting cells initiates the T H 1 fate, signals that initiate the T H 2 fate are not completely characterized. Here we show that early GATA-3 expression, required for T H 2 differentiation, was induced by T cell factor 1 (TCF-1) and its cofactor β-catenin, mainly from the proximal Gata3 promoter upstream of exon 1b. This activity was induced after T cell antigen receptor (TCR) stimulation and was independent of IL-4 receptor signaling through the transcription factor STAT6. Furthermore, TCF-1 blocked T H 1 fate by negatively regulating interferon-γ (IFN-γ) expression independently of β-catenin. Thus, TCF-1 initiates T H 2 differentiation of activated CD4 + T cells by promoting GATA-3 expression and suppressing IFN-γ expression. After being activated, naive CD4 + T cells differentiate into helper T cell subsets to orchestrate effective immunity to microorganisms. T helper type 1 (T H 1) cells mediate immunity against intracellular viruses and bacteria, whereas T H 2 cells protect against parasites and mediate allergic responses 1,2 . The best-characterized feature of helper T cells is that T H 1 cells produce interferon-γ (IFN-γ), whereas T H 2 cells produce interleukin 4 (IL-4; A001262), ). Whereas activated CD4 + T cells differentiate into the T H 1 lineage in response to IL-12 provided by antigen-presenting cells (APCs) and IFN-γ produced by natural killer cells 2 , similar signals that initiate T H 2 differentiation remain incompletely characterized. Once the T H 2 response is initiated, IL-4 production by activated CD4 + T cells reinforces the T H 2 fate5. However, the source of IL-4 at early stages of T H 2 differentiation remains Correspondence should be addressed to J.M.-S.

Role of Wnt Signaling in Colon Cancer Cell Metabolism

The FASEB Journal, Apr 1, 2016

Over eighty percent of colon cancers derive from mutations that overactivate the Wnt pathway and ... more Over eighty percent of colon cancers derive from mutations that overactivate the Wnt pathway and cause cell transformation. These mutations stabilize β-catenin protein, leading to its accumulation ...

Molecular Cancer Research, Mar 1, 2022

The recent classification of colon cancer into molecular subtypes revealed that patients with the... more The recent classification of colon cancer into molecular subtypes revealed that patients with the poorest prognosis harbor tumors with the lowest levels of Wnt signaling. This is contrary to the general understanding that overactive Wnt signaling promotes tumor progression from early initiation stages through to the later stages including invasion and metastasis. Here, we directly test this assumption by reducing the activity of ß-catenin-dependent Wnt signaling in colon cancer cell lines at either an upstream or downstream step in the pathway. We determine that Wntreduced cancer cells exhibit a more aggressive disease phenotype, including increased mobility in vitro and disruptive invasion into mucosa and smooth muscle in an orthotopic mouse model. RNA sequencing reveals that interference with Wnt signaling leads to an upregulation of gene programs that favor cell migration and invasion and a downregulation of inflammation signatures in the tumor microenvironment. We identify a set of upregulated genes common among the Wnt perturbations that are predictive of poor patient outcomes in early-invasive colon cancer. Our findings suggest that while targeting Wnt signaling may reduce tumor burden, an inadvertent side effect is the emergence of invasive cancer. Implications: Decreased Wnt signaling in colon tumors leads to a more aggressive disease phenotype due to an upregulation of gene programs favoring cell migration in the tumor and downregulation of inflammation programs in the tumor microenvironment; these impacts must be carefully considered in developing Wnt-targeting therapies.

The EMBO Journal, Aug 17, 2015

Cancer Biology & Therapy, 2008

Citing articles: 13 View citing articles b-catenin is the key transcriptional activator of the Wn... more Citing articles: 13 View citing articles b-catenin is the key transcriptional activator of the Wnt pathway important for development and tissue homeostasis of multicellular organisms. Its deregulation contributes to many human cancers. The b-catenin transcriptional activator complex continues to be defined, but already contains several proteins with chromatin remodeling activity. Here we show that two members of histone acetyltransferase complexes without enzymatic activity, hADA2a and hADA3, are required for full activity of b-catenin. hADA2a and hADA3 physically interact with b-catenin, and the interaction is mediated through Armadillo repeats 6 through 12 and the C-terminal transactivation domain of b-catenin. Both hADA2a and hADA3 reside with b-catenin at the enhancer for the Wnt target gene c-Myc. RNA interference-mediated reduction of hADA2a and hADA3 results in reduced b-catenin acetylation, reduced activity in reporter gene assays and reduced activation of endogenous b-catenin target genes. Overall, loss of hADA2a and hADA3 negatively impacts b-catenin-mediated proliferation. Our studies identify hADA2a and hADA3 as crucial cofactors of b-catenin that are likely involved in the assembly of transactivation-competent b-catenin complexes at Wnt target genes. b-catenin interacts with hADA2a and hADA3. hADA3 is known to interact directly with and be essential for three transcription factors with different cellular roles, p53, retinoic X receptor and estrogen receptor. We investigated whether hADA3 plays a role for b-catenin and included hADA2a because mouse ADA3

British Journal of Pharmacology, Aug 24, 2017

The highly conserved Wnt signalling pathway plays an important role in embryonic development and ... more The highly conserved Wnt signalling pathway plays an important role in embryonic development and disease pathogenesis, most notably cancer. The 'canonical' or β-catenin-dependent Wnt signal initiates at the cell plasma membrane with the binding of Wnt proteins to Frizzled:LRP5/LRP6 receptor complexes and is mediated by the translocation of the transcription co-activator protein, β-catenin, into the nucleus. β-Catenin then forms a complex with T-cell factor (TCF)/lymphoid enhancer binding factor (LEF) transcription factors to regulate multiple gene programmes. These programmes play roles in cell proliferation, migration, vasculogenesis, survival and metabolism. Mutations in Wnt signalling pathway components lead to constitutively active Wnt signalling that drives aberrant expression of these programmes and development of cancer. It has been a longstanding and challenging goal to develop therapies that can interfere with the TCF/LEF-β-catenin transcriptional complex. This review will focus on the (i) structural considerations for targeting the TCF/LEF-β-catenin and co-regulatory complexes in the nucleus, (ii) current molecules that directly target TCF/LEF-β-catenin activity and (iii) ideas for targeting newly discovered components of the TCF/LEF-β-catenin complex and/or downstream gene programmes regulated by these complexes.

Purification of TCF-1 alpha, a T-cell-specific transcription factor that activates the T-cell receptor C alpha gene enhancer in a context-dependent manner

PubMed, Jul 1, 1990

The differentiation of T cells into functionally diverse subpopulations is controlled in part, by... more The differentiation of T cells into functionally diverse subpopulations is controlled in part, by transcriptional activation and silencing; however, little is known in detail about the proteins that influence this developmental process. We have purified a new T-cell-specific factor, TCF-1 alpha, that is implicated in the activation of genes encoding a major component of the human T-cell receptor (TCR). TCF-1 alpha, originally identified and purified through its binding sites on the HIV-1 promoter, was found to bind to the TCR alpha enhancer and to promoters for several genes expressed at significantly earlier stages of T-cell development than the TCR alpha gene (e.g., p56lck and CD3 delta). Sequences related to the TCF-1 alpha binding motif (5'-GGCACCCTTTGA-3') are also found in the human TCR delta (and possibly TCR beta) enhancers. Southwestern and gel renaturation experiments with the use of purified protein fractions revealed that TCF-1 alpha activity is derived from a family of 57- to 53-kD proteins that are abundantly expressed in mature and immature T-cell lines (Jurkat, CCRF-CEM) and not in mature B cells (JY, Namalwa) or nonlymphoid (HeLa) cell lines. A small 95-bp fragment of the TCR alpha control region that contains the TCF-1 alpha binding site juxtaposed between a cAMP-response element (the CRE or T alpha 1 motif) and the binding site for a distinct lymphoid-specific protein (TCF-2 alpha) behaved as a potent T-cell-specific enhancer in vivo. Tandem copies of this enhancer functioned synergistically in mature (Jurkat) T-cell lines as well as resting and activated immature (CCRF-CEM) T-cell lines. Mutation of the TCF-1 alpha binding site diminished enhancer activity and disrupted the synergism observed in vivo between tandem enhancer repeats. The TCF-1 alpha binding site was also required for TCR alpha enhancer activity in transcriptionally active extracts from Jurkat but not HeLa cells, confirming that TCF-1 alpha is a T-cell-specific transcription factor. Curiously, the TCF-1 alpha binding element was inactive in vivo when removed from its neighboring elements on the TCR alpha enhancer and positioned in one or more copies upstream of a heterologous promoter. Thus, the transcriptional activity of TCF-1 alpha appears to depend on the TCF-2 alpha and T alpha 1 (CREB) transcription factors and the context of its binding site within the TCR alpha enhancer.

Journal of Biological Chemistry, May 1, 2003

Transcription of the lymphoid enhancer factor-1 (LEF1) gene is aberrantly activated in sporadic c... more Transcription of the lymphoid enhancer factor-1 (LEF1) gene is aberrantly activated in sporadic colon cancer, whereas this gene is not expressed in the normal adult colon. We have shown previously that promoter 1 of the LEF1 gene is activated by T cell factor (TCF)-␤catenin complexes in transient transfection assays, suggesting that LEF1 is a target of the Wnt pathway in colon cancer. To further explore the link between LEF1 expression and the Wnt pathway, we studied two response elements in the promoter. Surprisingly we found that the LEF1 promoter is selectively activated by specific isoforms of the LEF/TCF transcription factor family that contain an alternative C-terminal "E" tail. These isoforms, TCF-1E and TCF-4E, activate the promoter in a ␤-catenin-dependent manner. We show that a complete E-tail domain is necessary for full activity and delimits residues within two highly conserved peptide motifs within the tail that are required (KKCRARFG; WCXX-CRRKKKC). These peptide motifs are not only conserved among the TCF family members but are also found in two newly identified DNA-binding proteins named papillomavirus binding factor and GLUT4 enhancer factor. This study thus identifies a new and unique set of motifs used by the Wnt pathway for target gene regulation.

Molecular mechanisms of phorbol ester, thyrotropin-releasing hormone, and growth factor stimulation of prolactin gene transcription

Journal of Biological Chemistry, Sep 1, 1985

The polypeptide thyrotropin-releasing hormone (TRH) and epidermal growth factor (EGF) stimulate, ... more The polypeptide thyrotropin-releasing hormone (TRH) and epidermal growth factor (EGF) stimulate, within seconds to minutes, the transcription of the prolactin gene in a rat pituitary cell line (GH4). Because a series of agents that act to stimulate prolactin secretion fail to alter prolactin gene transcription, it is suggested that secretory events are neither obligatory for nor causal of hormone-induced transcriptional stimulation. Elevation of cytosolic-free calcium does not stimulate prolactin gene transcription; however, several agents that act to antagonize calcium-dependent processes inhibit or abolish both TRH and EGF stimulation of prolactin gene transcription and a specific hormone-dependent nuclear phosphorylation. In contrast, inhibitors of the slow calcium channel exert minimal effects on TRH-stimulated prolactin gene expression, suggesting that calcium influx through membrane channels is not crucial for the observed nuclear actions of TRH. Activation of protein kinase C by phorbol esters mimics the nuclear actions of TRH. In the presence of increased intracellular calcium levels, the effects of 12-O-tetradecanoyl phorbol 13-acetate on prolactin gene transcription are quantitatively identical to those observed in response to TRH or EGF.

Supplementary Data from Disruption of β-Catenin–Dependent Wnt Signaling in Colon Cancer Cells Remodels the Microenvironment to Promote Tumor Invasion

Supplementary Data from Disruption of β-Catenin–Dependent Wnt Signaling in Colon Cancer Cells Rem... more Supplementary Data from Disruption of β-Catenin–Dependent Wnt Signaling in Colon Cancer Cells Remodels the Microenvironment to Promote Tumor Invasion

Data from Disruption of β-Catenin–Dependent Wnt Signaling in Colon Cancer Cells Remodels the Microenvironment to Promote Tumor Invasion

The recent classification of colon cancer into molecular subtypes revealed that patients with the... more The recent classification of colon cancer into molecular subtypes revealed that patients with the poorest prognosis harbor tumors with the lowest levels of Wnt signaling. This is contrary to the general understanding that overactive Wnt signaling promotes tumor progression from early initiation stages through to the later stages including invasion and metastasis. Here, we directly test this assumption by reducing the activity of ß-catenin–dependent Wnt signaling in colon cancer cell lines at either an upstream or downstream step in the pathway. We determine that Wnt-reduced cancer cells exhibit a more aggressive disease phenotype, including increased mobility in vitro and disruptive invasion into mucosa and smooth muscle in an orthotopic mouse model. RNA sequencing reveals that interference with Wnt signaling leads to an upregulation of gene programs that favor cell migration and invasion and a downregulation of inflammation signatures in the tumor microenvironment. We identify a set of upregulated genes common among the Wnt perturbations that are predictive of poor patient outcomes in early-invasive colon cancer. Our findings suggest that while targeting Wnt signaling may reduce tumor burden, an inadvertent side effect is the emergence of invasive cancer.Implications:Decreased Wnt signaling in colon tumors leads to a more aggressive disease phenotype due to an upregulation of gene programs favoring cell migration in the tumor and downregulation of inflammation programs in the tumor microenvironment; these impacts must be carefully considered in developing Wnt-targeting therapies.Watch the interview with Marian L. Waterman, PhD, recipient of the 2023 MCR Michael B. Kastan Award for Research Excellence: https://vimeo.com/847435577

Role of Wnt Signaling in Colon Cancer Cell Metabolism

The FASEB Journal, Apr 1, 2016

Over eighty percent of colon cancers derive from mutations that overactivate the Wnt pathway and ... more Over eighty percent of colon cancers derive from mutations that overactivate the Wnt pathway and cause cell transformation. These mutations stabilize β-catenin protein, leading to its accumulation ...

Murdoch, G. H. , Waterman, M. , Evans, R. M. & Rosenfeld, M. G. Molecular mechanism of phorbol ester, thyrotropin-releasing hormone, and growth factor stimulation of prolactin gene transcription. J. Biol. Chem. 260, 11852-11858

Journal of Biological Chemistry

ABSTRACT

Feldman AS: Bladder cancer

Ring Finger Protein 14 is a new regulator of

Colorectal Cancer Stem Cell Subtypes Orchestrate Distinct Tumor Microenvironments

Nucleic Acids Research, May 1, 2000

Lymphoid Enhancer Factor-1 (LEF-1) is a member of a family of transcription factors that function... more Lymphoid Enhancer Factor-1 (LEF-1) is a member of a family of transcription factors that function as downstream mediators of the Wnt signal transduction pathway. In the absence of Wnt signals, specific LEF/TCF isoforms repress rather than activate gene targets through recruitment of the co-repressor CtBP. Characterization of the full-length human LEF-1 gene locus and its complete set of mRNA products shows that this family member exists as a unique set of alternatively spliced isoforms; none are homologous to TCF-1E/TCF-4E. Therefore LEF-1 is distinct from its TCF family members in that it cannot engage in activities specific to this isoform such as recruitment of the co-repressor CtBP. Expression of alternatively spliced LEF-1 isoforms are driven by a promoter that is highly active in lymphocyte cell lines. Transcription initiates within a TATA-less core promoter region that contains consensus binding sites for Sp1, an E box, an Initiator element and a LEF/TCF binding site, all juxtaposed to the start sites of transcription. The promoter is most active in a B lymphocyte cell line (Raji) in which the endogenous LEF-1 gene is silent, suggesting that the promoter region is actively repressed by a silencing mechanism.

Integrative Biology, 2014

Metastasis is the cause of over 90% of all human cancer deaths. Early steps in the metastatic pro... more Metastasis is the cause of over 90% of all human cancer deaths. Early steps in the metastatic process include: the formation of new blood vessels, the initiation of epithelial-mesenchymal transition (EMT), and the mobilization of tumor cells into the circulation. There are ongoing efforts to replicate the physiological landscape of human tumor tissue using three-dimensional in vitro culture models; however, few systems are able to capture the full range of authentic, complex in vivo events such as neovascularization and intravasation. Here we introduce the Prevascularized Tumor (PVT) model to investigate early events of solid tumor progression. PVT spheroids are composed of endothelial and tumor cells, and are embedded in a fibrin matrix containing fibroblasts. The PVT model facilitates two mechanisms of vessel formation: robust sprouting angiogenesis into the matrix, and contiguous vascularization within the spheroid. Furthermore, the PVT model enables the intravasation of tumor cells that is enhanced under low oxygen conditions and is also dependent on the key EMT transcription factor Slug. The PVT model provides a significant advance in the mimicry of human tumors in vitro and may improve investigation and targeting of events in the metastatic process.

Proceedings of the National Academy of Sciences of the United States of America, Apr 30, 2002

Life science alliance, Jul 13, 2022

Crosstalk between the Hedgehog and MAPK signaling pathways occurs in several types of cancer and ... more Crosstalk between the Hedgehog and MAPK signaling pathways occurs in several types of cancer and contributes to clinical resistance to Hedgehog pathway inhibitors. Here we show that MAP kinase-mediated phosphorylation weakens the binding of the GLI1 transcription factor to its negative regulator SUFU. ERK2 phosphorylates GLI1 on three evolutionarily conserved target sites (S102, S116, and S130) located near the high-affinity binding site for SUFU; these phosphorylations cooperate to weaken the affinity of GLI1-SUFU binding by over 25-fold. Phosphorylation of any one, or even any two, of the three sites does not result in the level of SUFU release seen when all three sites are phosphorylated. Tumor-derived mutations in R100 and S105, residues bordering S102, also diminish SUFU binding, collectively defining a novel evolutionarily conserved SUFU affinity-modulating region. In cultured mammalian cells, GLI1 variants containing phosphomimetic substitutions of S102, S116, and S130 displayed an increased ability to drive transcription. We conclude that multisite phosphorylation of GLI1 by ERK2 or other MAP kinases weakens GLI1-SUFU binding, thereby facilitating GLI1 activation and contributing to both physiological and pathological crosstalk.

Trends in Microbiology, 2008

Nature Immunology, Aug 2, 2009

The differentiation of activated CD4 + T cells into the T helper type 1 (T H 1) or T H 2 fate is ... more The differentiation of activated CD4 + T cells into the T helper type 1 (T H 1) or T H 2 fate is regulated by cytokines and the transcription factors T-bet and GATA-3. Whereas interleukin 12 (IL-12) produced by antigen-presenting cells initiates the T H 1 fate, signals that initiate the T H 2 fate are not completely characterized. Here we show that early GATA-3 expression, required for T H 2 differentiation, was induced by T cell factor 1 (TCF-1) and its cofactor β-catenin, mainly from the proximal Gata3 promoter upstream of exon 1b. This activity was induced after T cell antigen receptor (TCR) stimulation and was independent of IL-4 receptor signaling through the transcription factor STAT6. Furthermore, TCF-1 blocked T H 1 fate by negatively regulating interferon-γ (IFN-γ) expression independently of β-catenin. Thus, TCF-1 initiates T H 2 differentiation of activated CD4 + T cells by promoting GATA-3 expression and suppressing IFN-γ expression. After being activated, naive CD4 + T cells differentiate into helper T cell subsets to orchestrate effective immunity to microorganisms. T helper type 1 (T H 1) cells mediate immunity against intracellular viruses and bacteria, whereas T H 2 cells protect against parasites and mediate allergic responses 1,2 . The best-characterized feature of helper T cells is that T H 1 cells produce interferon-γ (IFN-γ), whereas T H 2 cells produce interleukin 4 (IL-4; A001262), ). Whereas activated CD4 + T cells differentiate into the T H 1 lineage in response to IL-12 provided by antigen-presenting cells (APCs) and IFN-γ produced by natural killer cells 2 , similar signals that initiate T H 2 differentiation remain incompletely characterized. Once the T H 2 response is initiated, IL-4 production by activated CD4 + T cells reinforces the T H 2 fate5. However, the source of IL-4 at early stages of T H 2 differentiation remains Correspondence should be addressed to J.M.-S.

Role of Wnt Signaling in Colon Cancer Cell Metabolism

The FASEB Journal, Apr 1, 2016

Over eighty percent of colon cancers derive from mutations that overactivate the Wnt pathway and ... more Over eighty percent of colon cancers derive from mutations that overactivate the Wnt pathway and cause cell transformation. These mutations stabilize β-catenin protein, leading to its accumulation ...

Molecular Cancer Research, Mar 1, 2022

The recent classification of colon cancer into molecular subtypes revealed that patients with the... more The recent classification of colon cancer into molecular subtypes revealed that patients with the poorest prognosis harbor tumors with the lowest levels of Wnt signaling. This is contrary to the general understanding that overactive Wnt signaling promotes tumor progression from early initiation stages through to the later stages including invasion and metastasis. Here, we directly test this assumption by reducing the activity of ß-catenin-dependent Wnt signaling in colon cancer cell lines at either an upstream or downstream step in the pathway. We determine that Wntreduced cancer cells exhibit a more aggressive disease phenotype, including increased mobility in vitro and disruptive invasion into mucosa and smooth muscle in an orthotopic mouse model. RNA sequencing reveals that interference with Wnt signaling leads to an upregulation of gene programs that favor cell migration and invasion and a downregulation of inflammation signatures in the tumor microenvironment. We identify a set of upregulated genes common among the Wnt perturbations that are predictive of poor patient outcomes in early-invasive colon cancer. Our findings suggest that while targeting Wnt signaling may reduce tumor burden, an inadvertent side effect is the emergence of invasive cancer. Implications: Decreased Wnt signaling in colon tumors leads to a more aggressive disease phenotype due to an upregulation of gene programs favoring cell migration in the tumor and downregulation of inflammation programs in the tumor microenvironment; these impacts must be carefully considered in developing Wnt-targeting therapies.

The EMBO Journal, Aug 17, 2015

Cancer Biology & Therapy, 2008

Citing articles: 13 View citing articles b-catenin is the key transcriptional activator of the Wn... more Citing articles: 13 View citing articles b-catenin is the key transcriptional activator of the Wnt pathway important for development and tissue homeostasis of multicellular organisms. Its deregulation contributes to many human cancers. The b-catenin transcriptional activator complex continues to be defined, but already contains several proteins with chromatin remodeling activity. Here we show that two members of histone acetyltransferase complexes without enzymatic activity, hADA2a and hADA3, are required for full activity of b-catenin. hADA2a and hADA3 physically interact with b-catenin, and the interaction is mediated through Armadillo repeats 6 through 12 and the C-terminal transactivation domain of b-catenin. Both hADA2a and hADA3 reside with b-catenin at the enhancer for the Wnt target gene c-Myc. RNA interference-mediated reduction of hADA2a and hADA3 results in reduced b-catenin acetylation, reduced activity in reporter gene assays and reduced activation of endogenous b-catenin target genes. Overall, loss of hADA2a and hADA3 negatively impacts b-catenin-mediated proliferation. Our studies identify hADA2a and hADA3 as crucial cofactors of b-catenin that are likely involved in the assembly of transactivation-competent b-catenin complexes at Wnt target genes. b-catenin interacts with hADA2a and hADA3. hADA3 is known to interact directly with and be essential for three transcription factors with different cellular roles, p53, retinoic X receptor and estrogen receptor. We investigated whether hADA3 plays a role for b-catenin and included hADA2a because mouse ADA3

British Journal of Pharmacology, Aug 24, 2017

The highly conserved Wnt signalling pathway plays an important role in embryonic development and ... more The highly conserved Wnt signalling pathway plays an important role in embryonic development and disease pathogenesis, most notably cancer. The 'canonical' or β-catenin-dependent Wnt signal initiates at the cell plasma membrane with the binding of Wnt proteins to Frizzled:LRP5/LRP6 receptor complexes and is mediated by the translocation of the transcription co-activator protein, β-catenin, into the nucleus. β-Catenin then forms a complex with T-cell factor (TCF)/lymphoid enhancer binding factor (LEF) transcription factors to regulate multiple gene programmes. These programmes play roles in cell proliferation, migration, vasculogenesis, survival and metabolism. Mutations in Wnt signalling pathway components lead to constitutively active Wnt signalling that drives aberrant expression of these programmes and development of cancer. It has been a longstanding and challenging goal to develop therapies that can interfere with the TCF/LEF-β-catenin transcriptional complex. This review will focus on the (i) structural considerations for targeting the TCF/LEF-β-catenin and co-regulatory complexes in the nucleus, (ii) current molecules that directly target TCF/LEF-β-catenin activity and (iii) ideas for targeting newly discovered components of the TCF/LEF-β-catenin complex and/or downstream gene programmes regulated by these complexes.

Purification of TCF-1 alpha, a T-cell-specific transcription factor that activates the T-cell receptor C alpha gene enhancer in a context-dependent manner

PubMed, Jul 1, 1990

The differentiation of T cells into functionally diverse subpopulations is controlled in part, by... more The differentiation of T cells into functionally diverse subpopulations is controlled in part, by transcriptional activation and silencing; however, little is known in detail about the proteins that influence this developmental process. We have purified a new T-cell-specific factor, TCF-1 alpha, that is implicated in the activation of genes encoding a major component of the human T-cell receptor (TCR). TCF-1 alpha, originally identified and purified through its binding sites on the HIV-1 promoter, was found to bind to the TCR alpha enhancer and to promoters for several genes expressed at significantly earlier stages of T-cell development than the TCR alpha gene (e.g., p56lck and CD3 delta). Sequences related to the TCF-1 alpha binding motif (5'-GGCACCCTTTGA-3') are also found in the human TCR delta (and possibly TCR beta) enhancers. Southwestern and gel renaturation experiments with the use of purified protein fractions revealed that TCF-1 alpha activity is derived from a family of 57- to 53-kD proteins that are abundantly expressed in mature and immature T-cell lines (Jurkat, CCRF-CEM) and not in mature B cells (JY, Namalwa) or nonlymphoid (HeLa) cell lines. A small 95-bp fragment of the TCR alpha control region that contains the TCF-1 alpha binding site juxtaposed between a cAMP-response element (the CRE or T alpha 1 motif) and the binding site for a distinct lymphoid-specific protein (TCF-2 alpha) behaved as a potent T-cell-specific enhancer in vivo. Tandem copies of this enhancer functioned synergistically in mature (Jurkat) T-cell lines as well as resting and activated immature (CCRF-CEM) T-cell lines. Mutation of the TCF-1 alpha binding site diminished enhancer activity and disrupted the synergism observed in vivo between tandem enhancer repeats. The TCF-1 alpha binding site was also required for TCR alpha enhancer activity in transcriptionally active extracts from Jurkat but not HeLa cells, confirming that TCF-1 alpha is a T-cell-specific transcription factor. Curiously, the TCF-1 alpha binding element was inactive in vivo when removed from its neighboring elements on the TCR alpha enhancer and positioned in one or more copies upstream of a heterologous promoter. Thus, the transcriptional activity of TCF-1 alpha appears to depend on the TCF-2 alpha and T alpha 1 (CREB) transcription factors and the context of its binding site within the TCR alpha enhancer.

Journal of Biological Chemistry, May 1, 2003

Transcription of the lymphoid enhancer factor-1 (LEF1) gene is aberrantly activated in sporadic c... more Transcription of the lymphoid enhancer factor-1 (LEF1) gene is aberrantly activated in sporadic colon cancer, whereas this gene is not expressed in the normal adult colon. We have shown previously that promoter 1 of the LEF1 gene is activated by T cell factor (TCF)-␤catenin complexes in transient transfection assays, suggesting that LEF1 is a target of the Wnt pathway in colon cancer. To further explore the link between LEF1 expression and the Wnt pathway, we studied two response elements in the promoter. Surprisingly we found that the LEF1 promoter is selectively activated by specific isoforms of the LEF/TCF transcription factor family that contain an alternative C-terminal "E" tail. These isoforms, TCF-1E and TCF-4E, activate the promoter in a ␤-catenin-dependent manner. We show that a complete E-tail domain is necessary for full activity and delimits residues within two highly conserved peptide motifs within the tail that are required (KKCRARFG; WCXX-CRRKKKC). These peptide motifs are not only conserved among the TCF family members but are also found in two newly identified DNA-binding proteins named papillomavirus binding factor and GLUT4 enhancer factor. This study thus identifies a new and unique set of motifs used by the Wnt pathway for target gene regulation.

Molecular mechanisms of phorbol ester, thyrotropin-releasing hormone, and growth factor stimulation of prolactin gene transcription

Journal of Biological Chemistry, Sep 1, 1985

The polypeptide thyrotropin-releasing hormone (TRH) and epidermal growth factor (EGF) stimulate, ... more The polypeptide thyrotropin-releasing hormone (TRH) and epidermal growth factor (EGF) stimulate, within seconds to minutes, the transcription of the prolactin gene in a rat pituitary cell line (GH4). Because a series of agents that act to stimulate prolactin secretion fail to alter prolactin gene transcription, it is suggested that secretory events are neither obligatory for nor causal of hormone-induced transcriptional stimulation. Elevation of cytosolic-free calcium does not stimulate prolactin gene transcription; however, several agents that act to antagonize calcium-dependent processes inhibit or abolish both TRH and EGF stimulation of prolactin gene transcription and a specific hormone-dependent nuclear phosphorylation. In contrast, inhibitors of the slow calcium channel exert minimal effects on TRH-stimulated prolactin gene expression, suggesting that calcium influx through membrane channels is not crucial for the observed nuclear actions of TRH. Activation of protein kinase C by phorbol esters mimics the nuclear actions of TRH. In the presence of increased intracellular calcium levels, the effects of 12-O-tetradecanoyl phorbol 13-acetate on prolactin gene transcription are quantitatively identical to those observed in response to TRH or EGF.

Supplementary Data from Disruption of β-Catenin–Dependent Wnt Signaling in Colon Cancer Cells Remodels the Microenvironment to Promote Tumor Invasion

Supplementary Data from Disruption of β-Catenin–Dependent Wnt Signaling in Colon Cancer Cells Rem... more Supplementary Data from Disruption of β-Catenin–Dependent Wnt Signaling in Colon Cancer Cells Remodels the Microenvironment to Promote Tumor Invasion

Data from Disruption of β-Catenin–Dependent Wnt Signaling in Colon Cancer Cells Remodels the Microenvironment to Promote Tumor Invasion

The recent classification of colon cancer into molecular subtypes revealed that patients with the... more The recent classification of colon cancer into molecular subtypes revealed that patients with the poorest prognosis harbor tumors with the lowest levels of Wnt signaling. This is contrary to the general understanding that overactive Wnt signaling promotes tumor progression from early initiation stages through to the later stages including invasion and metastasis. Here, we directly test this assumption by reducing the activity of ß-catenin–dependent Wnt signaling in colon cancer cell lines at either an upstream or downstream step in the pathway. We determine that Wnt-reduced cancer cells exhibit a more aggressive disease phenotype, including increased mobility in vitro and disruptive invasion into mucosa and smooth muscle in an orthotopic mouse model. RNA sequencing reveals that interference with Wnt signaling leads to an upregulation of gene programs that favor cell migration and invasion and a downregulation of inflammation signatures in the tumor microenvironment. We identify a set of upregulated genes common among the Wnt perturbations that are predictive of poor patient outcomes in early-invasive colon cancer. Our findings suggest that while targeting Wnt signaling may reduce tumor burden, an inadvertent side effect is the emergence of invasive cancer.Implications:Decreased Wnt signaling in colon tumors leads to a more aggressive disease phenotype due to an upregulation of gene programs favoring cell migration in the tumor and downregulation of inflammation programs in the tumor microenvironment; these impacts must be carefully considered in developing Wnt-targeting therapies.Watch the interview with Marian L. Waterman, PhD, recipient of the 2023 MCR Michael B. Kastan Award for Research Excellence: https://vimeo.com/847435577

Role of Wnt Signaling in Colon Cancer Cell Metabolism

The FASEB Journal, Apr 1, 2016

Over eighty percent of colon cancers derive from mutations that overactivate the Wnt pathway and ... more Over eighty percent of colon cancers derive from mutations that overactivate the Wnt pathway and cause cell transformation. These mutations stabilize β-catenin protein, leading to its accumulation ...

Murdoch, G. H. , Waterman, M. , Evans, R. M. & Rosenfeld, M. G. Molecular mechanism of phorbol ester, thyrotropin-releasing hormone, and growth factor stimulation of prolactin gene transcription. J. Biol. Chem. 260, 11852-11858

Journal of Biological Chemistry

ABSTRACT

Feldman AS: Bladder cancer

Ring Finger Protein 14 is a new regulator of

Colorectal Cancer Stem Cell Subtypes Orchestrate Distinct Tumor Microenvironments

Nucleic Acids Research, May 1, 2000

Lymphoid Enhancer Factor-1 (LEF-1) is a member of a family of transcription factors that function... more Lymphoid Enhancer Factor-1 (LEF-1) is a member of a family of transcription factors that function as downstream mediators of the Wnt signal transduction pathway. In the absence of Wnt signals, specific LEF/TCF isoforms repress rather than activate gene targets through recruitment of the co-repressor CtBP. Characterization of the full-length human LEF-1 gene locus and its complete set of mRNA products shows that this family member exists as a unique set of alternatively spliced isoforms; none are homologous to TCF-1E/TCF-4E. Therefore LEF-1 is distinct from its TCF family members in that it cannot engage in activities specific to this isoform such as recruitment of the co-repressor CtBP. Expression of alternatively spliced LEF-1 isoforms are driven by a promoter that is highly active in lymphocyte cell lines. Transcription initiates within a TATA-less core promoter region that contains consensus binding sites for Sp1, an E box, an Initiator element and a LEF/TCF binding site, all juxtaposed to the start sites of transcription. The promoter is most active in a B lymphocyte cell line (Raji) in which the endogenous LEF-1 gene is silent, suggesting that the promoter region is actively repressed by a silencing mechanism.

Integrative Biology, 2014

Metastasis is the cause of over 90% of all human cancer deaths. Early steps in the metastatic pro... more Metastasis is the cause of over 90% of all human cancer deaths. Early steps in the metastatic process include: the formation of new blood vessels, the initiation of epithelial-mesenchymal transition (EMT), and the mobilization of tumor cells into the circulation. There are ongoing efforts to replicate the physiological landscape of human tumor tissue using three-dimensional in vitro culture models; however, few systems are able to capture the full range of authentic, complex in vivo events such as neovascularization and intravasation. Here we introduce the Prevascularized Tumor (PVT) model to investigate early events of solid tumor progression. PVT spheroids are composed of endothelial and tumor cells, and are embedded in a fibrin matrix containing fibroblasts. The PVT model facilitates two mechanisms of vessel formation: robust sprouting angiogenesis into the matrix, and contiguous vascularization within the spheroid. Furthermore, the PVT model enables the intravasation of tumor cells that is enhanced under low oxygen conditions and is also dependent on the key EMT transcription factor Slug. The PVT model provides a significant advance in the mimicry of human tumors in vitro and may improve investigation and targeting of events in the metastatic process.