SHP-2 is required for the maintenance of cardiac progenitors

Langdon, Yvette; Goetz, Sarah C.; Berg, Anna E.; Swanik, Jackie Thomas; Conlon, Frank L.

doi:10.1242/dev.009290

Cited by 30 publications

(35 citation statements)

References 92 publications

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“…Whole-mount antibody staining was performed as described previously (Goetz et al, 2006;Langdon et al, 2007Langdon et al, , 2012 with mouse antitropomyosin (Developmental Studies Hybridoma Bank, 1:10) and Alexa 568 anti-mouse [1:250, IgG (H+L)]. Embryos were then cleared in 2:1 benzyl benzoate:benzyl alcohol and imaged on a Leica MZ16F fluorescence microscope with a Qimaging Retiga 4000RV Fast1394 camera with QCapture software as well as a Zeiss 700 confocal microscope with MyZen software.…”

Section: Immunohistochemistrymentioning

confidence: 99%

Congenital heart disease protein 5 associates with CASZ1 to maintain myocardial tissue integrity

et al. 2014

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The identification and characterization of the cellular and molecular pathways involved in the differentiation and morphogenesis of specific cell types of the developing heart are crucial to understanding the process of cardiac development and the pathology associated with human congenital heart disease. Here, we show that the cardiac transcription factor CASTOR (CASZ1) directly interacts with congenital heart disease 5 protein (CHD5), which is also known as tryptophanrich basic protein (WRB), a gene located on chromosome 21 in the proposed region responsible for congenital heart disease in individuals with Down's syndrome. We demonstrate that loss of CHD5 in Xenopus leads to compromised myocardial integrity, improper deposition of basement membrane, and a resultant failure of hearts to undergo cell movements associated with cardiac formation. We further report that CHD5 is essential for CASZ1 function and that the CHD5-CASZ1 interaction is necessary for cardiac morphogenesis. Collectively, these results establish a role for CHD5 and CASZ1 in the early stages of vertebrate cardiac development.

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Section: Immunohistochemistrymentioning

confidence: 99%

Congenital heart disease protein 5 associates with CASZ1 to maintain myocardial tissue integrity

et al. 2014

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“…two independent rounds of injections) except for stage 33, which was conducted four independent times. Results are reported as the ratio of triple to double positive cells ± two standard deviations by Student's t-test as previously described (Goetz et al, 2006;Langdon et al, 2007).…”

Section: Mitotic Index and Apoptosismentioning

confidence: 99%

“…All experiments on embryos were conducted as reported previously (Goetz et al, 2006;Langdon et al, 2007). Densitometry was used to standardize loading levels in western blot analysis.…”

Section: Western Blot Analysismentioning

confidence: 99%

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SHP-2 acts via ROCK to regulate the cardiac actin cytoskeleton

et al. 2012

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SUMMARYNoonan syndrome is one of the most common causes of human congenital heart disease and is frequently associated with missense mutations in the protein phosphatase SHP-2. Interestingly, patients with acute myelogenous leukemia (AML), acute lymphoblastic leukemia (ALL), juvenile myelomonocytic leukemia (JMML) and LEOPARD syndrome frequently carry a second, somatically introduced subset of missense mutations in SHP-2. To determine the cellular and molecular mechanisms by which SHP-2 regulates heart development and, thus, understand how Noonan-associated mutations affect cardiogenesis, we introduced SHP-2 encoding the most prevalent Noonan syndrome and JMML mutations into Xenopus embryos. Resulting embryos show a direct relationship between a Noonan SHP-2 mutation and its ability to cause cardiac defects in Xenopus; embryos expressing Noonan SHP-2 mutations exhibit morphologically abnormal hearts, whereas those expressing an SHP-2 JMML-associated mutation do not. Our studies indicate that the cardiac defects associated with the introduction of the Noonan-associated SHP-2 mutations are coupled with a delay or arrest of the cardiac cell cycle in M-phase and a failure of cardiomyocyte progenitors to incorporate into the developing heart. We show that these defects are a result of an underlying malformation in the formation and polarity of cardiac actin fibers and F-actin deposition. We show that these defects can be rescued in culture and in embryos through the inhibition of the Rho-associated, coiled-coil-containing protein kinase 1 (ROCK), thus demonstrating a direct relationship between SHP-2 N308D and ROCK activation in the developing heart.

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“…Furthermore, compared with other higher vertebrate models, Xenopus embryos can survive for longer without a properly functioning heart or circulatory system, enabling the observation of later consequences of cardiac malfunction. Moreover, the Xenopus model allows for tissue explantation for study in isolation (Afouda and Hoppler, 2009;Langdon et al, 2007;Raffin et al, 2000), as well as microinjection of overexpression constructs and antisense morpholino oligonucleotides to manipulate gene function (Brown et al, 2007;Brown et al, 2005;Christine and Conlon, 2008;Sive et al, 2000;Tada et al, 1997;Tandon et al, 2012). Crucially, studies have demonstrated a direct correlation between depleting cardiac proteins in Xenopus to that of genetic mutations in mouse or in human CHD patients (Kaltenbrun et al, 2011).…”

Section: Cardiac Profilingmentioning

confidence: 99%

Proteomic profiling of cardiac tissue by isolation of nuclei tagged in specific cell types (INTACT)

Amin

Greco

et al. 2014

Development

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The proper dissection of the molecular mechanisms governing the specification and differentiation of specific cell types requires isolation of pure cell populations from heterogeneous tissues and whole organisms. Here, we describe a method for purification of nuclei from defined cell or tissue types in vertebrate embryos using INTACT (isolation of nuclei tagged in specific cell types). This method, previously developed in plants, flies and worms, utilizes in vivo tagging of the nuclear envelope with biotin and the subsequent affinity purification of the labeled nuclei. In this study we successfully purified nuclei of cardiac and skeletal muscle from Xenopus using this strategy. We went on to demonstrate the utility of this approach by coupling the INTACT approach with liquid chromatography-tandem mass spectrometry (LC-MS/MS) proteomic methodologies to profile proteins expressed in the nuclei of developing hearts. From these studies we have identified the Xenopus orthologs of 12 human proteins encoded by genes, which when mutated in human lead to congenital heart disease. Thus, by combining these technologies we are able to identify tissue-specific proteins that are expressed and required for normal vertebrate organ development.

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SHP-2 is required for the maintenance of cardiac progenitors

Cited by 30 publications

References 92 publications

Congenital heart disease protein 5 associates with CASZ1 to maintain myocardial tissue integrity

Congenital heart disease protein 5 associates with CASZ1 to maintain myocardial tissue integrity

SHP-2 acts via ROCK to regulate the cardiac actin cytoskeleton

Proteomic profiling of cardiac tissue by isolation of nuclei tagged in specific cell types (INTACT)

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