-
Donna Lai,
Xifu Liu, Ariel Forrai,
Orit Wolstein,
Jan Michalicek,
Ishtiaq Ahmed,
Alistair N Garratt,
Carmen Birchmeier,
Mingdong Zhou,
Lynne Hartley,
Lorraine Robb,
Michael P Feneley,
Diane Fatkin,
Richard P Harvey
[show abstract]
[hide abstract]
ABSTRACT: The cardiac gene regulatory network (GRN) is controlled by transcription factors and signaling inputs, but network logic in development and it unraveling in disease is poorly understood. In development, the membrane-tethered signaling ligand Neuregulin (Nrg)1, expressed in endocardium, is essential for ventricular morphogenesis. In adults, Nrg1 protects against heart failure and can induce cardiomyocytes to divide.
To understand the role of Nrg1 in heart development through analysis of null and hypomorphic Nrg1 mutant mice.
Chamber domains were correctly specified in Nrg1 mutants, although chamber-restricted genes Hand1 and Cited1 failed to be activated. The chamber GRN subsequently decayed with individual genes exhibiting decay patterns unrelated to known patterning boundaries. Both trabecular and nontrabecular myocardium were affected. Network demise was spatiotemporally dynamic, the most sensitive region being the central part of the left ventricle, in which the GRN underwent complete collapse. Other regions were partially affected with graded sensitivity. In vitro, Nrg1 promoted phospho-Erk1/2-dependent transcription factor expression, cardiomyocyte maturation and cell cycle inhibition. We monitored cardiac pErk1/2 in embryos and found that expression was Nrg1-dependent and levels correlated with cardiac GRN sensitivity in mutants.
The chamber GRN is fundamentally labile and dependent on signaling from extracardiac sources. Nrg1-ErbB1/4-Erk1/2 signaling critically sustains elements of the GRN in trabecular and nontrabecular myocardium, challenging our understanding of Nrg1 function. Transcriptional decay patterns induced by reduced Nrg1 suggest a novel mechanism for cardiac transcriptional regulation and dysfunction in disease, potentially linking biomechanical feedback to molecular pathways for growth and differentiation.
Circulation Research 09/2010; 107(6):715-27. · 9.49 Impact Factor
-
Ariel Forrai,
Kristy Boyle,
Adam H Hart,
Lynne Hartley,
Steven Rakar,
Tracy A Willson,
Ken M Simpson,
Andrew W Roberts,
Warren S Alexander,
Anne K Voss,
Lorraine Robb
[show abstract]
[hide abstract]
ABSTRACT: Leukemia inhibitory factor (LIF) is required to maintain pluripotency and permit self-renewal of murine embryonic stem (ES) cells. LIF binds to a receptor complex of LIFR-beta and gp130 and signals via the Janus kinase-signal transducer and activator of transcription (JAK-STAT) pathway, with signalling attenuated by suppressor of cytokine signalling (SOCS) proteins. Recent in vivo studies have highlighted the role of SOCS-3 in the negative regulation of signalling via gp130. To determine the role of SOCS-3 in ES cell biology, SOCS-3-null ES cell lines were generated. When cultured in LIF levels that sustain self-renewal of wild-type cells, SOCS-3-null ES cell lines exhibited less self-renewal and greater differentiation into primitive endoderm. The absence of SOCS-3 enhanced JAK-STAT and extracellular signal-related kinase 1/2 (ERK-1/2)-mitogen-activated protein kinase (MAPK) signal transduction via gp130, with higher levels of phosphorylated STAT-1, STAT-3, SH-2 domain-containing cytoplasmic protein tyrosine phosphatase 2 (SHP-2), and ERK-1/2 in steady state and in response to LIF stimulation. Attenuation of ERK signalling by the addition of MAPK/ERK kinase (MEK) inhibitors to SOCS-3-null ES cell cultures rescued the differentiation phenotype, but did not restore proliferation to wild-type levels. In summary, SOCS-3 plays a crucial role in the regulation of the LIF signalling pathway in murine ES cells. Its absence perturbs the balance between activation of the JAK-STAT and SHP-2-ERK-1/2-MAPK pathways, resulting in less self-renewal and a greater potential for differentiation into the primitive endoderm lineage.
Stem Cells 04/2006; 24(3):604-14. · 7.78 Impact Factor
-
[show abstract]
[hide abstract]
ABSTRACT: The laboratory mouse is an invaluable tool for functional gene discovery because of its genetic malleability and a biological similarity to human systems that facilitates identification of human models of disease. A number of mutagenic technologies are being used to elucidate gene function in the mouse. Gene trapping is an insertional mutagenesis strategy that is being undertaken by multiple research groups, both academic and private, in an effort to introduce mutations across the mouse genome. Large-scale, publicly funded gene trap programs have been initiated in several countries with the International Gene Trap Consortium coordinating certain efforts and resources. We outline the methodology of mammalian gene trapping and how it can be used to identify genes expressed in both primitive and definitive blood cells and to discover hemopoietic regulator genes. Mouse mutants with hematopoietic phenotypes derived using gene trapping are described. The efforts of the large-scale gene trapping consortia have now led to the availability of libraries of mutagenized ES cell clones. The identity of the trapped locus in each of these clones can be identified by sequence-based searching via the world wide web. This resource provides an extraordinary tool for all researchers wishing to use mouse genetics to understand gene function.
Experimental Hematology 09/2005; 33(8):845-56. · 2.90 Impact Factor
-
Ariel Forrai,
Kristy Boyle,
Adam H. Hart,
Lynne Hartley,
Steven Rakar,
Tracy A. Willson,
Ken M. Simpson,
Andrew W. Roberts,
Warren S. Alexander,
Anne K. Voss,
Ph.D. Lorraine Robb M.D
[show abstract]
[hide abstract]
ABSTRACT: Leukemia inhibitory factor (LIF) is required to maintain pluripotency and permit self-renewal of murine embryonic stem (ES) cells. LIF binds to a receptor complex of LIFR-β and gp130 and signals via the Janus kinase–signal transducer and activator of transcription (JAK–STAT) pathway, with signalling attenuated by suppressor of cytokine signalling (SOCS) proteins. Recent in vivo studies have highlighted the role of SOCS-3 in the negative regulation of signalling via gp130. To determine the role of SOCS-3 in ES cell biology, SOCS-3–null ES cell lines were generated. When cultured in LIF levels that sustain self-renewal of wild-type cells, SOCS-3–null ES cell lines exhibited less self-renewal and greater differentiation into primitive endoderm. The absence of SOCS-3 enhanced JAK–STAT and extracellular signal–related kinase 1/2 (ERK-1/2)–mitogen-activated protein kinase (MAPK) signal transduction via gp130, with higher levels of phosphorylated STAT-1, STAT-3, SH-2 domain–containing cytoplasmic protein tyrosine phosphatase 2 (SHP-2), and ERK-1/2 in steady state and in response to LIF stimulation. Attenuation of ERK signalling by the addition of MAPK/ERK kinase (MEK) inhibitors to SOCS-3–null ES cell cultures rescued the differentiation phenotype, but did not restore proliferation to wild-type levels. In summary, SOCS-3 plays a crucial role in the regulation of the LIF signalling pathway in murine ES cells. Its absence perturbs the balance between activation of the JAK–STAT and SHP-2–ERK-1/2–MAPK pathways, resulting in less self-renewal and a greater potential for differentiation into the primitive endoderm lineage.
Stem Cells 08/2005; 24(3):604 - 614. · 7.78 Impact Factor
-
[show abstract]
[hide abstract]
ABSTRACT: The hemangioblast is a bipotential cell that gives rise to hematopoietic and endothelial cells. Although the existence of the hemangioblast was first postulated early last century, a cell with this activity has yet to be unequivocally identified in mammals. In the last decade, gene targeting experiments in the mouse have uncovered genes which are required for development of both the hematopoietic and endothelial lineages, and this, together with increasing recognition that the two cell types share gene expression patterns, has renewed interest in the hemangioblast. The murine embryonic stem cell differentiation system has been used to demonstrate the existence of a Fft-1 positive progenitor cell, called the BL-CFC, which has the properties of the hemangioblast and this system is now being used to dissect the molecular regulation of hemangioblast development and differentiation.
Cell cycle (Georgetown, Tex.) 2(2):86-90. · 5.36 Impact Factor
