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Regulatory Strategies of the Neuroendocrine System in Development, Physiology, and Disease
Summary: Michael Rosenfeld is investigating integrative nuclear strategies responsible for orchestrating programs of genome-wide transcriptional responses to diverse signaling systems, including the endocrine system, that are critical for physiological and behavioral processes in all vertebrates. His work continues to reveal unexpected gene-specific strategies that link regulated gene responses to other cellular response programs, including DNA damage/repair and development. Defining these strategies has suggested new approaches to diseases, including growth defects, diabetes, atherosclerosis, and several prevalent forms of cancer.
Recent studies have revealed that intrinsic enzymatic activities in a diverse network of coactivator and corepressor complexes—acting as "sensors" to permit integration of multiple signaling pathways—modulate, reorganize, and fine-tune patterns of transcriptional response. Unexpectedly, hormonal regulation of gene expression includes transient double-stranded DNA cleavage as a required component of regulated gene transcription. It also includes noncoding RNAs that act as ligands by regulating specific gene activation and repression programs. Definition of a complex coregulatory network has included definition of combinatorial codes for regulation of specific cohorts of gene targets. New genome-wide approaches have uncovered a series of novel mechanisms that permit the complex transcriptional response programs required for all metazoans. Elucidating these programs has provided insights into development and disease.
Methylation-Dependent Mechanisms and Ligand Dependency
Nuclear receptors undergo ligand-dependent conformational changes required for corepressor-coactivator exchange, but whether gene activation requires specific epigenetic landmarks to impose ligand dependency remains unknown. Recently we uncovered an unexpected and general strategy that is based on the requirement for specific cohorts of inhibitory histone methyltransferases (HMTs) to impose gene-specific "gatekeeper" functions. These functions prevent unliganded nuclear receptors and other classes of regulated transcription factors from binding to their target gene promoters and causing constitutive gene activation in the absence of stimulating signals. This strategy is based, in part, on an HMT-dependent inhibitory histone code, as well as methylation of other regulatory targets. It imposes a requirement for specific histone demethylases, including LSD1, to permit ligand- and signal-dependent activation of regulated gene expression. These events link an inhibitory methylation activity, including histone substrates, to a broadly used strategy that circumvents pathological constitutive gene induction by physiologically regulated transcription factors.
Opposing LSD1 Complexes in Gene Activation and Repression Programs in the Neuroendocrine System
Precise control of transcriptional programs underlying metazoan development is modulated by enzymatically active coregulatory complexes, coupled with epigenetic strategies. How specific members of histone modification enzyme families, such as histone methyltransferases and demethylases, are utilized in vivo to orchestrate distinct developmental gene activation and repression programs remains unclear. We used a conditional gene deletion strategy to investigate functions of LSD1 in development of the anterior pituitary gland, a well-understood model of organogenesis. We found that LSD1, a component of the CoREST/CtBP-containing corepressor complex, is required for late cell-lineage determination and differentiation during pituitary organogenesis. Surprisingly, LSD1 acts primarily on target gene activation programs, as well as in gene repression programs, based on recruitment of distinct LSD1-containing coactivator or corepressor complexes. LSD1-dependent gene repression programs can be extended late in development with the induced expression of ZEB1, a Krüpple-like repressor that can act as a molecular beacon for recruitment of the LSD1-containing CtBP/CoREST corepressor complex. This represses an additional cohort of genes, such as GH, that previously required LSD1 for activation.
LSD1 thus regulates specific developmental programs following initial organ commitment and prior to cell-type differentiation. Initially LSD1 serves as a key component of opposing coactivator and corepressor complexes. These complexes, which are recruited in a gene-specific fashion, are required for dictating specific programs of gene expression during mammalian organogenesis. It is likely that LSD1 activates a cohort of gene targets, including GH (growth hormone) gene expression, by removing repressive histone marks, but nonhistone substrates are suggested to be biologically important. The temporally regulated, cell-type-specific restriction of GH gene expression in the emerging lactotrope population after birth is apparently based, at least in part, on postpartum estrogen-dependent induction of ZEB1. This Krüpple-like zinc finger DNA-binding repressor, and at least two other components of the CtBP/CoREST/LSD1 corepressor complex, together assemble an LSD1-containing corepressor complex on the GH promoter. Thus, after birth, LSD1 is present on the GH gene promoter in a coactivator complex in cells expressing GH (somatotropes), where it is important in GH gene activation, and in a ZEB1-recruited corepressor complex on the GH promoter in lactotropes in which the GH gene is repressed.
Signaling Pathways in Development
Although the roles of canonical Wnt/β-catenin signaling in development and disease are well documented, understanding the molecular logic underlying the nuclear transcriptional programs that mediate the diverse functions of β-catenin is a major challenge. Analysis of pituitary organogenesis has revealed an unexpected strategy for β-catenin-dependent regulation of cell-lineage determination, based on interactions between a β-catenin and a specific homeodomain factor, Prop1, rather than LEF/TCFs. β-Catenin acts as a binary switch to activate simultaneous expression of the critical lineage-determining transcription factor, Pit1, and repression of the gene encoding the lineage-inhibiting transcription factor, Hesx1, with β-catenin acting via TLE/Reptin/HDAC1 corepressors. The functionally distinct actions of a homeodomain factor in response to Wnt signaling are suggested to be prototypic of a widely used mechanism for generating diverse cell types from pluripotent precursor cells in response to common signaling pathways during organogenesis.
Sustained Notch activity has proved to be required for the temporal maintenance of specific cohorts of proliferating precursors in the developing pituitary gland, which underlies the ability to specify late-arising cell lineages. Conditional deletion of the major mediator of the Notch pathway, CSL/Rbp-J, leads to premature differentiation of progenitor cells and conversion of the late (Pit1) lineage into the early (corticotrope) lineage. Sustained Notch signaling in progenitor cells is thus required to prevent conversion of the late-arising cell lineages to early-born cell lineages, permitting specification of diverse cell types in mammalian organogenesis.
Macrophage-Cancer Cell Interactions
Androgens, acting via androgen receptors, are essential for growth of the normal prostate gland and prostate cancer. Although selective androgen receptor antagonists/modulators (SARMs) are initially effective in treatment of prostate cancer, there is rapid invariant resistance, with progression from androgen-dependent to androgen-independent growth.
Our studies have revealed that a subset of NFκB-regulated genes, including a key metastasis-suppressor gene, KAI1, can be derepressed in response to proinflammatory cytokines produced by activated macrophages. Based on these findings, we identified robust direct interactions between macrophages and prostate cell lines: virtually 100 percent of samples in prostate cancer tissue arrays exhibit specific macrophage–cancer cell interactions. These interactions mediate a switch in SARM function from repression to activation in vivo. This switch is based on the selective presence of an evolutionarily conserved receptor amino-terminal helical motif in sex steroid receptors that modulates recruitment of a factor (TAB2) as a component of the N-CoR corepressor complex. TAB2 serves as a molecular beacon for recruitment of protein kinase MEKK1, which mediates dismissal of the N-CoR complex, causing derepression of androgen and estrogen receptor (AR and ER) target genes. Peptides corresponding to the helical motif of AR or ERα receptors can block macrophage-dependent resistance. This suggests that nonpeptidergic orthologs might prevent inflammatory cytokine-dependent switch in SARM or SERM function and hence "block" resistance. Liganded SUMOylated nuclear receptors, including PPARγ and LXR, are recruited to NCoR/SMRT corepressor complexes to transrepress gene programs activated by inflammatory cytokines, based on blocking recruitment of Ubc5 ubiquitylation machinery required for dismissal of the corepressor complex. This provides potential new therapeutic approaches for treatment of prostate cancer.
These studies were supported in part by grants from the National Institutes of Health and the National Cancer Institute.
Last updated: March 16, 2007
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