1999 — 2001 |
Sucov, Henry M |
R01Activity Code Description: To support a discrete, specified, circumscribed project to be performed by the named investigator(s) in an area representing his or her specific interest and competencies. |
Pathway of Ventricular Morphogenesis Medicated by Rxra @ University of Southern California
In a surprisingly large number of mouse gene knockout studies, a common phenotype has been observed which involves a failure of fetal ventricular cardiomyocytes to proliferate during development and form a thickened outer ventricular chamber wall (the "compact zone"). These genes are the retinoic acid receptor (and nuclear transcription factor) RXRalpha, the cell cycle regulator N-myc, the transcription factors TEF1 and WT1, the cell surface receptor GP130, the cell surface receptor kinase (betaARK1, and the cell adhesion molecules VCAM1 and alpha4 integrin. Little is known of the manner in which most of the above-mentioned genes act. The assignment of these genes which regulate compact zone cardiomyocyte proliferation to their tissue of action and to pathways of action will define the nature of the events which underlie cardiac ventricular morphogenesis. Specific Aim 1: Define the cell autonomy of the TEF1, WT1, GP13O, and betaARK1 mutant phenotypes. New data indicates that RXRalpha functions outside of the cardiomyocyte lineage, and other investigators have determined the site of action of N-myc, alpha4 integrin, and VCAM1. By derivation of embryonic stem cells lacking TEF1, WT1, GP130, or betaARK1, and by evaluation of the behavior of these cells in chimeric embryos, it will be possible to determine if these genes act within or outside the cardiomyocyte lineage. Specific Aim 2: Evaluate candidate tissues as the site of action of RXRalpha. A combination of transgenic and chimeric approaches will define the tissue in which RXRalpha function is required for normal cardiomyocyte proliferation in the ventricular chamber wall. Specific Aim 3: Evaluate candidate tissues as the site of action of other genes which act in a cell nonautonomous manner. Transgenic and chimeric approaches will be employed to address the site of action of TEF1, WT1, GP130, and betaARK1 (if these act outside of the cardiomyocyte lineage), and to define pathways which unite the deficiency of these gene products into a common mutant phenotype. A greater understanding of the manner in which the genes described in this proposal act might shed light not only on the etiology of certain congenital heart defects, but also on general mechanisms which regulate the distinction between atrial and ventricular morphogenesis (the atrial chamber not undergoing formation of a compact zone), and on the loss of proliferative competence in postnatal cardiomyocytes.
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0.958 |
2000 — 2001 |
Sucov, Henry M |
R21Activity Code Description: To encourage the development of new research activities in categorical program areas. (Support generally is restricted in level of support and in time.) |
Prostate Development Using Cre Recombinase @ University of Southern California
A new transgenic mouse model (called PB-cre), in which cre recombinase is expressed specifically in the male prostate under the control of the prostate-specific probasin promoter, has been derived for the study of prostate development and cancer. In this R2l application, we first propose to characterize the expression pattern of this transgene and to define the onset and extent of recombination over time of conditional genes. We then propose to selectively mutate the retinoid receptor RXR alpha: gene in prostatic epithelium in normal mice and in mice genetically disposed to prostatic epithelial dysplasia. Specific Aim 1: To define the pattern of recombination driven by the PB- cre transgene. A comprehensive description of the timing and extent of recombination as driven by this transgene will be obtained by crossing to the well-characterized conditional reporter gene ROSA26R (R26R). Specific Aim 2: To selectively mutate the RXR alpha: gene in prostate epithelium. Abundant prior and new evidence presented in this proposal implicate a role for retinoids and retinoid receptors, particularly RXR alpha:, in the derivation and maintenance of prostatic epithelium and in the progression of prostate cancer. We will cross the crc-expressing transgene with a conditional allele of RXR alpha:, and evaluate the nature and extent of morphological alterations in the prostatic epithelium. We will also cross the conditional mutation of RXR alpha: into the Nkx3. l knockout mouse model to evaluate the extent to which RXR alpha: modulates the initiation and progression of prostate dysplasia and cancer.
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0.958 |
2001 — 2004 |
Sucov, Henry M |
R01Activity Code Description: To support a discrete, specified, circumscribed project to be performed by the named investigator(s) in an area representing his or her specific interest and competencies. |
Retinoic Acid Regulation of Erythropoiesis @ University of Southern California
The cytokine erythropoietin (Epo) initiates proliferation and differentiation along the red blood cell lineage. The Epo gene is physiologically regulated by hypoxia, through a defined element in the Epo gene 3' enhancer. Epo expression additionally requires factors which bind to a small direct repeat sequence known as a DR2 element, which is adjacent to the hypoxia responsive site. DR2 elements serve as binding sites for members of the nuclear receptor family, which includes HNF4 and retinoic acid (RA) receptors. While HNF4 has previously been implicated as a regulator of Epo expression through the DR2 sequence, there has been no prior indication that the Epo gene is RA responsive. New evidence derived from analysis of knockout mice and through molecular and biochemical approaches suggests that retinoic acid is in fact a regulator of Epo gene expression in the fetal liver, and thereby regulates definitive erythropoiesis. Hypothesis and model: In early definitive hematopoiesis, erythropoietic differentiation is regulated by retinoic acid and the retinoic acid receptor RXRalpha by direct transcriptional activation of Epo gene expression through the Epo enhancer DR2. In later erythropoiesis, there is a transition to regulation by HNF4. Specific Aim 1: Demonstrate that Epo gene expression is directly regulated by retinoic acid and by RXRalpha. Extensive preliminary data presented in this proposal indicate that the Epo gene is a downstream target gene of retinoic acid action. In this Aim, I will confirm that the Epo gene is directly activated by retinoic acid, and that the nature of this regulation occurs through recognition by RA receptors of the Epo enhancer DR2 element. Specific Aim 2: Define the molecular basis of the onset of erythropoiesis in RXRalpha-/- embryos. In this Aim, I will test the hypothesis that a transition occurs in definitive erythropoiesis from regulation of Epo gene expression retinoic acid and RXRalpha to regulation by HNF4.
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0.958 |
2002 — 2006 |
Sucov, Henry M |
R01Activity Code Description: To support a discrete, specified, circumscribed project to be performed by the named investigator(s) in an area representing his or her specific interest and competencies. |
Cardiomyocyte Proliferation &Ventricular Morphogenesis @ University of Southern California
DESCRIPTION (provided by applicant): The mechanisms which promote cardiomyocyte proliferation during fetal life, and which fail to induce proliferation during postnatal life, are only poorly understood. My lab has studied cardiac development in mouse embryos lacking the retinoic acid receptor RXRa, in which compact zone formation in the ventricular chamber fails to initiate, resulting in a hypoplastic chamber that is incompatible with fetal viability. We have previously described this phenotype and shown that the initial response to retinoic acid during ventricular development does not occur in the myocardium, but rather occurs in a distinct cell lineage which then secondarily supports myocardial proliferation and subsequent ventricular chamber morphogenesis. Model: Based on new data presented in this application, I propose that retinoic acid signaling is initially transduced by RXRa in the epicardium, thereby inducing expression of one or more secreted growth factors which are trophic for the myocardium. Myocardium in older embryos and in postnatal animals is refractory to stimulation by these factors. To address this model, I propose the following Specific Aims: Specific Aim 1: To define the role of RXRa in the epicardium. Tissue-specific inactivation and transgenic manipulation in mouse embryos, and viral infection in chick embryos, are proposed to demonstrate that RXRa function occurs in the epicardium. Specific Aim 2: To identify the factor(s) secreted by epicardial cells in response to retinoic acid that are trophic for the myocardium. Biochemical purification plus gene-based discovery approaches are proposed to identify the factors secreted by epicardial cells in a retinoic acid- and RXRa-dependent manner. Specific Aim 3: To define the critical components of the myocardial response to epicardial cell secreted trophic factor(s). I propose to identify the signal transduction elements in fetal cardiocytes which mediate the response to trophic stimulation, as well as changes which account for the lack of response in postnatal myocardium.
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0.958 |
2006 — 2009 |
Sucov, Henry M |
R01Activity Code Description: To support a discrete, specified, circumscribed project to be performed by the named investigator(s) in an area representing his or her specific interest and competencies. |
Tgfb and Cardiac Neural Crest Cell Fate and Function @ University of Southern California
DESCRIPTION (provided by applicant): This project addresses signaling processes during formation of the aorta and pulmonary trunk, and then in the maturation and maintenance of these vessels. A greater understanding of these processes might lead to diagnostic or therapeutic opportunities for malformations that occur when these processes go awry. We have undertaken neural crest-specific disruption of the type II TGFb receptor gene (Tgfbr2) in mice, to address the role of TGFb signaling in the formation and maturation of the outflow tract vessels of the developing heart. Mutant embryos have persistent truncus arteriosus (PTA) and interrupted aortic arch (IAA- B);later in gestation, the elastic matrix of the neural crest-derived vessel wall of the ductus arteriosus becomes disorganized, leading to vessel wall dilation (aneurysm). The central premise to explain these defects is that the behavior of neural crest cells is dictated by local signaling events, that neural crest cells respond directly to these signals, and that TGFb is one such signal. Specific Aim 1: To identify mechanisms by which absence of TGFb signaling in neural crest cells results in PTA. In this Aim, we will identify and characterize the role of genes that are misregulated in neural crest- derived smooth muscle cells;we will also address the requirement for Smad-dependent signaling pathways downstream of the TGFb receptor. Specific Aim 2: To address the role of retinoic acid and RA receptors in the formation of the A/P septum, and the potential convergence of this pathway with TGFb signaling pathways. Mutation of retinoic acid receptor genes leads to a virtually identical PTA defect as in Tgfbr2 mutants. We will define the tissue in which retinoic acid signaling occurs, and address the convergence of RA and TGFb signaling pathways in the process of outflow tract septation. Specific Aim 3: To address the role of TGFb signaling in vessel wall maturation. In this Aim, we will study the onset of vascular wall deformation, address the extent to which TGFb signal transduction is required in mesodermally-derived vs. neural crest-derived smooth muscle and resolve whether vessel dilation results directly or indirectly from a mechanically impaired vessel wall.
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0.958 |
2009 — 2019 |
Sucov, Henry M |
R01Activity Code Description: To support a discrete, specified, circumscribed project to be performed by the named investigator(s) in an area representing his or her specific interest and competencies. |
Cardiomyocyte Proliferation and Ventricular Morphogenesis @ University of Southern California
DESCRIPTION (provided by applicant): This project addresses mechanisms that are responsible for formation of the ventricular chamber of the heart during mouse embryonic development. We have advanced a model in which signals secreted from the epicardium promote cell division and morphological organization of the underlying compact zone myocardium. In this application, we propose to pursue the following goals: Specific Aim 1: To define the role of IGF signaling in heart development. Several lines of evidence presented in this proposal indicate that IGF signaling is an important mediator of proliferation in the midgestation heart, although a functional role for IGF signaling in heart development has not been examined previously. In this Aim, we will primarily undertake a phenotypic assessment of mouse embryos in which IGF ligands and receptors are conventionally and conditionally mutated. Specific Aim 2: To define pathways that lead to morphogenic differentiation of the compact and trabecular myocardium under the influence of the epicardium and endocardium. Epicardial and endocardial factors converge on a common set of signaling intermediates to activate proliferation, yet diverge to achieve differential morphogenic outcomes. How the ventricular myocardium becomes partitioned into compact and trabecular compartments is a critical aspect of heart development, yet one that is not well- understood. In this Aim, we address the molecular pathways that result in the selective expression of genes in the compact zone or trabecular myocardium, with the consideration that these are surrogate markers for the larger biological processes of compact vs. trabecular morphogenesis. PUBLIC HEALTH RELEVANCE: This project considers how the muscle of the embryonic heart forms and becomes organized. An understanding of these processes may give insight into certain forms of congenital heart defects, and may be relevant to therapies to reverse or treat the pathology of adult heart failure.
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0.958 |
2015 — 2016 |
Sucov, Henry M |
R21Activity Code Description: To encourage the development of new research activities in categorical program areas. (Support generally is restricted in level of support and in time.) |
Cardiomyocyte Heterogeneity and Aging @ University of Southern California
? DESCRIPTION (provided by applicant): This project seeks to define the complexity of cardiomyocyte populations in the adult mouse ventricle, and has a specific emphasis on understanding the nature of proliferative cardiomyocytes that support adult heart regeneration after injury. We begin from the perspective that proliferative cardiomyocytes in the adult mouse heart are mononuclear and diploid, which corresponds to the proliferative population in mouse neonatal and zebrafish adult hearts and to the proliferative population in the embryonic hearts of all species. We propose to overcome several outstanding issues related to this general conceptualization that have impeded progress in the field. First, the extent to which the adult mononuclear diploid cardiomyocyte population is heterogeneous is unknown, particularly as related to subpopulations that have proliferative competence. Second, no molecular markers currently exist that identify proliferative cardiomyocytes from nonproliferative. Third, the dynamics of these populations in the heart over time are completely unknown, but are likely of critical importance in understanding the prevalence of heart failure in the elderly. In Aim 1, we will undertake single cell RNA Seq analysis of mononuclear diploid, mononuclear tetraploid, and binuclear cardiomyocytes isolated from adult mice, and will show that genes or gene isoforms that are differentially expressed define groups and subpopulations of these groups of cardiomyocytes. This approach will demonstrate the extent of cardiomyocyte heterogeneity and at the same time will validate markers of these populations that can be used in a number of subsequent studies. As one particularly important application, we will identify which markers label proliferative cardiomyocytes after injury. In Aim 2, we will address the dynamics of these populations over time, in particular comparing young adult to aged adult mice. We will resolve among several models to explain the observation that cardiomyocyte regeneration declines with advanced age. In all, this project combines an innovative conceptualization with new technology to address questions about the substructure and dynamics of cardiomyocyte populations that have not previously been accessible.
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0.958 |
2019 — 2021 |
Sucov, Henry M |
R01Activity Code Description: To support a discrete, specified, circumscribed project to be performed by the named investigator(s) in an area representing his or her specific interest and competencies. |
The Complex Genetics of Heart Regeneration @ Medical University of South Carolina
Project Summary Shortly after birth, most mammalian cardiomyocytes (CMs) become postmitotic and nonregenerative, concurrent with becoming polyploid (either binucleated or mononuclear tetraploid). What controls this process has been unknown. As a result, though, it is generally thought that the adult heart has too little regenerative capacity to appreciably recover after injury. A subpopulation of mononuclear diploid CMs (MNDCMs), thought to be very small in number, persists in the heart through adulthood and is a candidate cell type to support endogenous heart regeneration. Using a wholly new conceptualization, we demonstrated in inbred mice that the percentage of MNDCMs in the adult heart, and the degree of functional recovery and of CM proliferation after adult heart injury, are both highly variable traits subject to the combined influence of multiple polymorphic genes. We identified and confirmed the CM- specific kinase Tnni3k as one such gene with polymorphic alleles that influence variability in the adult level of MNDCMs and thereby in the level of CM regeneration after adult infarction. Using this new understanding, the focus of this project is to elucidate the processes that cause CMs to become polyploid and postmitotic, and the consequences of these processes. In Aim 1, we will dissociate the two roles of Tnni3k in the natural neonatal heart after birth and in the adult heart following injury; we propose that these are related but independent and both relevant to the outcome after adult injury. In Aim 2, we propose to identify two new genes that also influence how CMs remain proliferative or become postmitotic, and to confirm our insights related to the regenerative capacity of MNDCMs. In Aim 3, we explore an unexpected influence of Tnni3k in the proper function of the cardiac conduction system (the electrical system of the heart), and how this role relates to the process of heart regeneration. In Aim 4, we invoke a mechanistic understanding to unify these observations of CM regeneration and conduction. The conceptual significance of this project is transformational: rather than heart injury resulting inexorably in declining heart function as is currently believed, some individuals may have substantial regenerative ability depending on their genetic and cellular composition. Furthermore, these insights might be developed for therapeutics to improve heart regeneration in all patients regardless of their genetic background. And finally, this project might explain why the mammalian heart transitions in most individuals to become mostly postmitotic in the postnatal period, even at the cost of loss of regenerative ability.
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0.925 |