Serrate and Notch specify cell fates in the heart field by suppressing cardiomyogenesis

Notch signaling mediates numerous developmental cell fate decisions in organisms ranging from flies to humans, resulting in the generation of multiple cell types from equipotential precursors. In this paper, we present evidence that activation of Notch by its ligand Serrate apportions myogenic and non-myogenic cell fates within the early Xenopus heart field. The crescent-shaped field of heart mesoderm is specified initially as cardiomyogenic. While the ventral region of the field forms the myocardial tube, the dorsolateral portions lose myogenic potency and form the dorsal mesocardium and pericardial roof (Raffin, M., Leong, L. M., Rones, M. S., Sparrow, D., Mohun, T. and Mercola, M. (2000) Dev. Biol., 218, 326–340). The local interactions that establish or maintain the distinct myocardial and non-myocardial domains have never been described. Here we show that Xenopus Notch1 (Xotch) and Serrate1 are expressed in overlapping patterns in the early heart field. Conditional activation or inhibition of the Notch pathway with inducible dominant negative or active forms of the RBP-J/Suppressor of Hairless [Su(H)] transcription factor indicated that activation of Notch feeds back on Serrate1 gene expression to localize transcripts more dorsolaterally than those of Notch1, with overlap in the region of the developing mesocardium. Moreover, Notch pathway activation decreased myocardial gene expression and increased expression of a marker of the mesocardium and pericardial roof, whereas inhibition of Notch signaling had the opposite effect. Activation or inhibition of Notch also regulated contribution of individual cells to the myocardium. Importantly, expression of Nkx2. 5 and Gata4 remained largely unaffected, indicating that Notch signaling functions downstream of heart field specification. We conclude that Notch signaling through Su(H) suppresses cardiomyogenesis and that this activity is essential for the correct specification of myocardial and non-myocardial cell fates.

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Abstract 3380: Acetylation-dependent Regulation Of Notch1 Signaling In Endothelial Cells

Circulation ◽

10.1161/circ.118.suppl_18.s_415-c ◽

2008 ◽

Vol 118 (suppl_18) ◽

Author(s):

Virginia Guarani ◽

Franck Dequiedt ◽

Andreas M Zeiher ◽

Stefanie Dimmeler ◽

Michael Potente

Keyword(s):

Endothelial Cells ◽

Notch Signaling ◽

Cell Fate ◽

Molecular Mechanisms ◽

Trichostatin A ◽

Notch Pathway ◽

Dependent Manner ◽

Class Iii ◽

Cell Fate Decisions ◽

Knock Down

The Notch signaling pathway is a versatile regulator of cell fate decisions and plays an essential role for embryonic and postnatal vascular development. As only modest differences in Notch pathway activity suffice to determine dramatic differences in blood vessel development, this pathway is tightly regulated by a variety of molecular mechanisms. Reversible acetylation has emerged as an important post-translational modification of several non-histone proteins, which are targeted by histone deacetylases (HDACs). Here, we report that specifically the Notch1 intracellular domain (NICD) is itself an acetylated protein and that its acetylation level is tightly regulated by the SIRT1 deacetylase, which we have previously identified as a key regulator of endothelial angiogenic functions during vascular growth. Coexpression of NICD with histone acetyltransferases such as p300 or PCAF induced a dose- and time-dependent acetylation of NICD. Blocking HDAC activity using the class III HDAC inhibitor nicotinamid (NAM), but not the class I/II HDAC inhibior trichostatin A, resulted in a significant increase of NICD acetylation suggesting that NICD is targetd by class III HDACs for deacetylation. Among the class III HDACs with deacetylase activity (SIRT1, 2, 3, 5), knock down of specifically SIRT1 resulted in enhanced acetylation of NICD. Moreover, wild type SIRT1, but not a catalytically inactive mutant catalyzed the deacetylation of NICD in a nicotinamid-dependent manner. SIRT1, but SIRT2, SIRT3 or SIRT5, associated with NICD through its catalytic domain demonstrating that SIRT1 is a direct NICD deacetylase. Enhancing NICD acetylation by either overexpression of p300 or inhibition of SIRT1 activity using NAM or RNAi-mediated knock down resulted in enhanced NICD protein stability by blocking its ubiquitin-mediated degradation. Consistent with these results, loss of SIRT1 amplified Notch target gene expression in endothelial cells in response to NICD overexpression or treatment with the Notch ligand Dll4. In summary, our results identify reversible acetylation of NICD as a novel molecular mechanism to control Notch signaling and suggest that deacetylation of NICD by SIRT1 plays a key role in the dynamic regulation of Notch signaling in endothelial cells.

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Hypoxia-Inducible Factors 1α and 2α Regulate Trophoblast Differentiation

Molecular and Cellular Biology ◽

10.1128/mcb.25.23.10479-10491.2005 ◽

2005 ◽

Vol 25 (23) ◽

pp. 10479-10491 ◽

Cited By ~ 132

Author(s):

Karen D. Cowden Dahl ◽

Benjamin H. Fryer ◽

Fiona A. Mack ◽

Veerle Compernolle ◽

Emin Maltepe ◽

...

Keyword(s):

Cell Fate ◽

Cell Types ◽

Hypoxia Inducible Factors ◽

Low Oxygen ◽

Placental Development ◽

Cell Fates ◽

Trophoblast Stem Cells ◽

Cell Fate Decisions ◽

Hypoxic Environment ◽

Life Threatening

ABSTRACT Placental development initially occurs in a low-oxygen (O2) or hypoxic environment. In this report we show that two hypoxia-inducible factors (HIFs), HIF1α and HIF2α, are essential for determining murine placental cell fates. HIF is a heterodimer composed of HIFα and HIFβ (ARNT) subunits. Placentas from Arnt − / − and Hif1α − / − Hif2α −/− embryos exhibit defective placental vascularization and aberrant cell fate adoption. HIF regulation of Mash2 promotes spongiotrophoblast differentiation, a prerequisite for trophoblast giant cell differentiation. In the absence of Arnt or Hifα, trophoblast stem cells fail to generate these cell types and become labyrinthine trophoblasts instead. Therefore, HIF mediates placental morphogenesis, angiogenesis, and cell fate decisions, demonstrating that O2 tension is a critical regulator of trophoblast lineage determination. This novel genetic approach provides new insights into the role of O2 tension in the development of life-threatening pregnancy-related diseases such as preeclampsia.

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Cancer Stem Cells, Quo Vadis? The Notch Signaling Pathway in Tumor Initiation and Progression

Cells ◽

10.3390/cells9081879 ◽

2020 ◽

Vol 9 (8) ◽

pp. 1879 ◽

Cited By ~ 2

Author(s):

Christian T. Meisel ◽

Cristina Porcheri ◽

Thimios A. Mitsiadis

Keyword(s):

Stem Cells ◽

Cancer Stem Cells ◽

Notch Signaling ◽

Signaling Pathway ◽

Cell Fate ◽

Adult Life ◽

Notch Pathway ◽

Notch Signaling Pathway ◽

Fine Tuning ◽

Cell Fate Decisions

The Notch signaling pathway regulates cell proliferation, cytodifferentiation and cell fate decisions in both embryonic and adult life. Several aspects of stem cell maintenance are dependent from the functionality and fine tuning of the Notch pathway. In cancer, Notch is specifically involved in preserving self-renewal and amplification of cancer stem cells, supporting the formation, spread and recurrence of the tumor. As the function of Notch signaling is context dependent, we here provide an overview of its activity in a variety of tumors, focusing mostly on its role in the maintenance of the undifferentiated subset of cancer cells. Finally, we analyze the potential of molecules of the Notch pathway as diagnostic and therapeutic tools against the various cancers.

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Foxi1 inactivation rescues loss of principal cell fate selection in Hes1-deficient kidneys but does not ensure maintenance of principal cell gene expression

10.1101/826610 ◽

2019 ◽

Author(s):

Malini Mukherjee ◽

Jennifer DeRiso ◽

Madhusudhana Janga ◽

Eric Fogarty ◽

Kameswaran Surendran

Keyword(s):

Gene Expression ◽

Notch Signaling ◽

Cell Fate ◽

Collecting Duct ◽

Cell Types ◽

Principal Cell ◽

Intercalated Cell ◽

Collecting Ducts ◽

Cell Gene Expression ◽

Cell Gene

AbstractThe distal nephron and collecting duct segments of the mammalian kidney consist of intercalated cell types intermingled among principal cell types. Notch signaling ensures that a sufficient number of cells select a principal instead of an intercalated cell fate. However, the precise mechanisms by which Notch signaling patterns the distal nephron and collecting duct cell fates is unknown. Here we observed that Hes1, a direct target of Notch signaling pathway, is required within the mouse developing collecting ducts for repression of Foxi1 expression, an essential intercalated cell specific transcription factor. Interestingly, inactivation of Foxi1 in Hes1-deficient collecting ducts rescues the deficiency in principal cell fate selection, overall urine concentrating deficiency, and reduces the occurrence of hydronephrosis. However, Foxi1 inactivation does not rescue the reduction in expression of all principal cell genes in the Hes1-deficient kidney collecting duct cells that select the principal cell fate. Additionally, suppression of Notch/Hes1 signaling in mature principal cells reduces principal cell gene expression without activating Foxi1. We conclude that Hes1 is a Notch signaling target that is essential for normal patterning of the collecting ducts with intermingled cell types by repressing Foxi1, and for maintenance of principal cell gene expression independent of repressing Foxi1.

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Notch1 inhibits differentiation of hematopoietic cells by sustaining GATA-2 expression

Blood ◽

10.1182/blood.v98.12.3283 ◽

2001 ◽

Vol 98 (12) ◽

pp. 3283-3289 ◽

Cited By ~ 96

Author(s):

Keiki Kumano ◽

Shigeru Chiba ◽

Kiyoshi Shimizu ◽

Tetsuya Yamagata ◽

Noriko Hosoya ◽

...

Keyword(s):

Transcription Factors ◽

Notch Signaling ◽

Cell Fate ◽

Hematopoietic Cells ◽

Erythroid Differentiation ◽

Inhibitory Effect ◽

Dominant Negative ◽

Granulocytic Differentiation ◽

Cell Fate Decisions ◽

Delayed Differentiation

Abstract Notch signaling is involved in cell fate decisions in many systems including hematopoiesis. It has been shown that expression of an activated form of Notch1 (aNotch1) in 32D mouse myeloid progenitor cells inhibits the granulocytic differentiation induced by granulocyte colony-stimulating factor (G-CSF). Results of the current study show that aNotch1, when expressed in F5-5 mouse erythroleukemia cells, also inhibits erythroid differentiation. Comparison of the expression levels of several transcription factors after stimulation for myeloid and erythroid differentiation, in the presence or absence of aNotch1, revealed that aNotch1 did not change its regulation pattern with any of the transcription factors examined, except for GATA-2, despite its inhibitory effect on differentiation. GATA-2 was down-regulated when the parental 32D and F5-5 were induced to differentiate into granulocytic and erythroid lineages, respectively. In these induction procedures, however, the level of GATA-2 expression was sustained when aNotch1 was expressed. To ascertain whether maintenance of GATA-2 is required for the Notch-induced inhibition of differentiation, the dominant-negative form of GATA-3 (DN-GATA), which acted also against GATA-2, or transcription factor PU.1, which was recently shown to be the repressor of GATA-2, was introduced into aNotch1-expressing 32D (32D/aNotch1) cells that do not express GATA family proteins other than GATA2. Both DN-GATA and PU.1 reversed the phenotype of 32D/aNotch1 inducing its differentiation when G-CSF was added. Furthermore, enforced expression of HES-1, which is involved in Notch signaling, delayed differentiation of 32D, and again this phenotype was neutralized by DN-GATA. These results indicate that GATA-2 activity is necessary for the Notch signaling in hematopoietic cells.

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The Drosophila melanogaster Suppressor of deltex Gene, a Regulator of the Notch Receptor Signaling Pathway, Is an E3 Class Ubiquitin Ligase

Genetics ◽

10.1093/genetics/152.2.567 ◽

1999 ◽

Vol 152 (2) ◽

pp. 567-576 ◽

Cited By ~ 13

Author(s):

M Cornell ◽

D A P Evans ◽

R Mann ◽

M Fostier ◽

M Flasza ◽

...

Keyword(s):

Notch Signaling ◽

Signaling Pathway ◽

Cell Fate ◽

Ubiquitin Ligase ◽

Notch Pathway ◽

Notch Receptor ◽

Negative Regulator ◽

Loss Of Function ◽

Wing Vein ◽

Cell Fate Decisions

Abstract During development, the Notch receptor regulates many cell fate decisions by a signaling pathway that has been conserved during evolution. One positive regulator of Notch is Deltex, a cytoplasmic, zinc finger domain protein, which binds to the intracellular domain of Notch. Phenotypes resulting from mutations in deltex resemble loss-of-function Notch phenotypes and are suppressed by the mutation Suppressor of deltex [Su(dx)]. Homozygous Su(dx) mutations result in wing-vein phenotypes and interact genetically with Notch pathway genes. We have previously defined Su(dx) genetically as a negative regulator of Notch signaling. Here we present the molecular identification of the Su(dx) gene product. Su(dx) belongs to a family of E3 ubiquitin ligase proteins containing membrane-targeting C2 domains and WW domains that mediate protein-protein interactions through recognition of proline-rich peptide sequences. We have identified a seven-codon deletion in a Su(dx) mutant allele and we show that expression of Su(dx) cDNA rescues Su(dx) mutant phenotypes. Overexpression of Su(dx) also results in ectopic vein differentiation, wing margin loss, and wing growth phenotypes and enhances the phenotypes of loss-of-function mutations in Notch, evidence that supports the conclusion that Su(dx) has a role in the downregulation of Notch signaling.

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Unique functions for Notch4 in murine embryonic lymphangiogenesis

10.1101/2021.02.19.431982 ◽

2021 ◽

Author(s):

Ajit Muley ◽

Minji Kim Uh ◽

Jennifer M. James ◽

Aino Murtomaki ◽

Joseph D. McCarron ◽

...

Keyword(s):

Endothelial Cell ◽

Notch Signaling ◽

Cell Fate ◽

Dominant Role ◽

Lymphatic Endothelial Cell ◽

Dominant Negative ◽

Endothelial Cell Migration ◽

Endothelial Cell Proliferation ◽

Lymphatic Endothelium ◽

Cell Fate Decisions

AbstractIn mice, embryonic dermal lymphatic development is a well-understood system used to study the role of genes in physiological lymphangiogenesis. The Notch signaling is an evolutionary conserved pathway that modulates cell fate decisions and shown to both inhibit and promote dermal lymphangiogenesis. Here, we demonstrate distinct roles for Notch4 signaling versus canonical Notch signaling in embryonic dermal lymphangiogenesis. At E14.5, actively growing dermal lymphatics expressed NOTCH1, NOTCH4 and DLL4, with DLL4 expression strongest and Notch active in the lymphangiogenic sprouts. Treatment of cultured LECs with VEGF-A or VEGF-C upregulated Dll4 transcripts, but differentially regulated Notch1 and Notch4 expression, and the Notch effectors of the Hes/Hey families, suggesting that VEGF-A and VEGF-C distinctly modulate Dll4/Notch signaling in the lymphatic endothelium. Mice nullizygous for Notch4 had an increase in the closure of the lymphangiogenic fronts towards the midline which correlated with reduced vessel caliber in the maturing lymphatic plexus. Activation of Notch4 suppressed lymphatic endothelial cell migration in a wounding assay significantly more then Notch1 activation, suggesting a dominant role for Notch4 in LEC migration. Unlike Notch4 nulls, inhibition of canonical Notch signaling by ectopically expressing a dominant negative form of MAML1 (DNMAML) in Prox1+ lymphatic endothelium suppressed lymphatic endothelial cell proliferation consistent with what has been described for the loss of lymphatic endothelial Notch1. Moreover, loss of Notch4 did not disrupt lymphatic endothelial canonical Notch signaling. Thus, we propose that Notch4 signaling and canonical Notch signaling have distinct functions in the coordination of embryonic dermal lymphangiogenesis.

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Mastermind critically regulates Notch-mediated lymphoid cell fate decisions

Blood ◽

10.1182/blood-2004-02-0514 ◽

2004 ◽

Vol 104 (6) ◽

pp. 1696-1702 ◽

Cited By ~ 194

Author(s):

Ivan Maillard ◽

Andrew P. Weng ◽

Andrea C. Carpenter ◽

Carlos G. Rodriguez ◽

Hong Sai ◽

...

Keyword(s):

T Cell ◽

B Cell ◽

Cell Fate ◽

Critical Role ◽

Family Members ◽

Notch Pathway ◽

Dominant Negative ◽

Hematopoietic Stem ◽

Cell Fate Decisions

Abstract During lymphoid development, Notch1 plays a critical role in the T-cell/B-cell lineage decision, while Notch2 is essential for marginal zone B-cell (MZB) development. Notch pathway activation induces translocation of intracellular Notch (ICN) to the nucleus, where it interacts with the transcription factor CSL (CBF1/RBP-Jk, Suppressor of Hairless, Lag-1). In vitro, ICN binds Mastermind-like proteins, which act as potent Notch coactivators. Three MAML family members (MAML1-3) have been identified in mammals, but their importance in vivo is unknown. To investigate the function of MAMLs in hematopoietic development, we introduced a dominant negative (DN) mutant of MAML1, capable of inhibiting Notch1-4, in murine hematopoietic stem cells. DNMAML1 resulted in early inhibition of T-cell development and the appearance of intrathymic B cells, phenotypes consistent with Notch1 inhibition. The T-cell differentiation block was as profound as that produced by enforced expression of the Notch modulator Deltex1. In DNMAML1-transduced spleen cells, a dramatic decrease in MZB cells was present, consistent with Notch2 inhibition. In contrast, Deltex1 did not decrease MZB cell numbers. These results suggest a critical role for MAMLs during Notch-mediated cell fate decisions in vivo and indicate that DNMAML1, but not Deltex1, can be used to interfere with the function of multiple Notch family members. (Blood. 2004;104:1696-1702)

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Notch Signaling Induces Megakaryocytic Cell Fate.

Blood ◽

10.1182/blood.v110.11.200.200 ◽

2007 ◽

Vol 110 (11) ◽

pp. 200-200

Author(s):

Thomas Mercher ◽

Melanie Cornejo ◽

Christopher Sears ◽

Thomas Kindler ◽

Sandra Moore ◽

...

Keyword(s):

Bone Marrow ◽

Bone Marrow Transplantation ◽

Notch Signaling ◽

Cell Fate ◽

Marrow Transplantation ◽

Notch Pathway ◽

Severe Thrombocytopenia ◽

Hematopoietic Stem ◽

Cell Fate Decisions

Abstract The Notch pathway regulates a broad range of biological mechanisms including proliferation, border formation and cell fate decisions. In the hematopoietic system, Notch signaling is generally thought to specify T cell lineage fate at the expense of the B cell whereas its role in the myeloid lineage development is unclear. When using heterotypic co-cultures of murine primary hematopoietic stem cells (HSC: Lin-Sca1+Kit+) with OP9 stromal cells, or OP9 cells expressing the Notch ligand Delta1 (OP9-DL1), we unexpectedly observed the development of large cells with cytoplasmic protrusions reminiscent of proplatelet production by megakaryocytes on OP9-DL1 stroma. These cells stained positive for acetylcholinesterase, specific for megakaryocyte, and displayed large polylobated nuclei. Flow cytometric analysis indicated a 10-fold increase in the number of CD41+ cells in OP9-DL1 co-cultures compared to parental OP9 co-cultures. Expression of a constitutively active intra-cellular Notch (ICN) mutant allowed differentiation of HSC into CD41+ cells in parental OP9 co-cultures without DL1 stimulation, whereas expression of a dominant-negative MAML1 (dnMAML1) mutant abrogated this effect in OP9-DL1 co-cultures. In addition, megakaryocyte differentiation in OP9-DL1 co-cultures was blocked by γ-secretase inhibitors treatment and rescued by ectopic expression of ICN. Global gene expression analysis demonstrated engagement of a megakaryopoietic transcriptional program when HSC were co-cultured with OP9-DL1 vs. OP9 stroma or OP9-DL1 stroma treated with γ-secretase inhibitor. Bone marrow transplantation experiments with ICN, resulted in enhanced megakaryopoiesis in vivo with increased MEP numbers and megakaryocyte colony formation. Furthermore, transplantation of bone marrow cells transduced with dnMAML1 significantly impaired megakaryopoiesis in vivo with a 4- to 7-fold decrease in maturing megakaryocytes. These findings demonstrate a positive regulatory role for Notch signaling in specification of megakaryocyte development, and indicate that Notch plays a complex role in cell fate decisions among myeloid progenitors. They suggest the possibility that inhibition of Notch signaling may have therapeutic potential in malignancies of the megakaryocytic lineage. Furthermore, Notch pathway stimulation could be of value in enhancing megakaryocyte growth in clinical contexts associated with severe thrombocytopenia, such as hematopoietic reconstitution following bone marrow transplantation or chemotherapy.

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Differential gene expression in a coculture model of angiogenesis reveals modulation of select pathways and a role for Notch signaling

Physiological Genomics ◽

10.1152/physiolgenomics.90318.2008 ◽

2009 ◽

Vol 36 (2) ◽

pp. 69-78 ◽

Cited By ~ 33

Author(s):

Brenda Lilly ◽

Simone Kennard

Keyword(s):

Gene Expression ◽

Endothelial Cells ◽

Notch Signaling ◽

Three Dimensional ◽

Notch Pathway ◽

Cell Types ◽

Three Dimensional Model ◽

Mural Cells ◽

Expression Of Genes ◽

Coculture Model

Communication between endothelial and mural cells (smooth muscle cells, pericytes, and fibroblasts) can dictate blood vessel size and shape during angiogenesis, and control the functional aspects of mature blood vessels, by determining things such as contractile properties. The ability of these different cell types to regulate each other's activities led us to ask how their interactions directly modulate gene expression. To address this, we utilized a three-dimensional model of angiogenesis and screened for genes whose expression was altered under coculture conditions. Using a BeadChip array, we identified 323 genes that were uniquely regulated when endothelial cells and mural cells (fibroblasts) were cultured together. Data mining tools revealed that differential expression of genes from the integrin, blood coagulation, and angiogenesis pathways were overrepresented in coculture conditions. Scans of the promoters of these differentially modulated genes identified a multitude of conserved C promoter binding factor (CBF)1/CSL elements, implicating Notch signaling in their regulation. Accordingly, inhibition of the Notch pathway with γ-secretase inhibitor DAPT or NOTCH3-specific small interfering RNA blocked the coculture-induced regulation of several of these genes in fibroblasts. These data show that coculturing of endothelial cells and fibroblasts causes profound changes in gene expression and suggest that Notch signaling is a critical mediator of the resultant transcription.

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