Our long-term objectives are to elucidate the cellular, molecular and genetic mechanisms that regulate early myocardial cell differentiation and myofibrillogenesis in developing hearts. A naturally occurring recessive lethal mutation in axolotls (salamanders), Ambystoma mexicanum, is an intriguing model for studying early heart development, because homozygous embryos (c/c) form hearts that are deficient in tropomyosin, lack organized myofibrils, and fail to beat. The defect can be corrected by culturing hearts with normal anterior endoderm tissue, in medium conditioned by the anterior endoderm, or in total RNA isolated from endoderm or conditioned medium. In addition, we have identified a novel Clone (#4) from a cDNA library constructed from conditioned medium RNA which acts as a template for a bioactive RNA capable of correcting the heart defect. The RNA can bind at least two proteins in the embryos. It is possible that this RNA is a regulatory molecule which upregulates tropomyosin synthesis in mutant hearts and promotes myofibrillogenesis either directly or in complex with its binding protein(s). The following specific aims have been developed to address our hypothesis that this RNA, most likely in conjunction with RNA binding proteins, promotes myocardial cell differentiation and myofibrillogenesis in developing heart cells: 1) The role of the Clone #4 RNA in promoting myofibrillogenesis in embryonic hearts will be determined; 2) Transgenic axolotls will be used to test the ability of Clone #4 to rescue mutant hearts in vivo; 3) Expression of the bioactive Clone #4 RNA will be analyzed in developing embryos by in situ hybridization and quantitative RT-PCR; 4) The structural-functional relationships of the bioactive Clone #4 RNA will be evaluated by in vitro mutagenesis; 5) The proteins that bind to the bioactive RNA will be purified and characterized at the cellular, biochemical and molecular levels; 6) Potential homolog(s) of the bioactive Clone #4 RNA will be examined in other animal systems. This research will provide new basic information on the molecular mechanisms Of myofibrillogenesis and myocardial cell rescue in vertebrate hearts. The health relevance of understanding and being able to turn defective, non-beating cardiac tissue into contracting muscle could be tremendous; if this can be applied in humans, patients who have damaged heart tissue might be able to have that tissue redifferentiate into functional muscle again.

Agency
National Institute of Health (NIH)
Institute
National Heart, Lung, and Blood Institute (NHLBI)
Type
Research Project (R01)
Project #
2R01HL061246-06A1
Application #
6576496
Study Section
Special Emphasis Panel (ZRG1-CVB (02))
Program Officer
Schramm, Charlene A
Project Start
1998-03-15
Project End
2007-03-31
Budget Start
2003-04-01
Budget End
2004-03-31
Support Year
6
Fiscal Year
2003
Total Cost
$242,389
Indirect Cost
Name
Florida Atlantic University
Department
Type
Other Domestic Higher Education
DUNS #
004147534
City
Boca Raton
State
FL
Country
United States
Zip Code
33431
Kochegarov, Andrei; Moses-Arms, Ashley; Lemanski, Larry F (2015) A fetal human heart cardiac-inducing RNA (CIR) promotes the differentiation of stem cells into cardiomyocytes. In Vitro Cell Dev Biol Anim 51:739-48
Moses-Arms, Ashley; Kochegarov, Andrei; Arms, Jedidiah et al. (2015) Identification of a human mitochondrial RNA that promotes tropomyosin synthesis and myocardial differentiation. In Vitro Cell Dev Biol Anim 51:273-80
Kochegarov, Andrei; Moses, Ashley; Lian, William et al. (2013) A new unique form of microRNA from human heart, microRNA-499c, promotes myofibril formation and rescues cardiac development in mutant axolotl embryos. J Biomed Sci 20:20
Jia, Pingping; Zhang, Chi; Jia, Yuanyuan et al. (2011) Identification of a truncated form of Methionine Sulfoxide Reductase A expressed in mouse embryonic stem cells. J Biomed Sci 18:46
Zhang, Chi; Jia, Pingping; Jia, Yuanyuan et al. (2011) Anoxia, acidosis, and intergenic interactions selectively regulate methionine sulfoxide reductase transcriptions in mouse embryonic stem cells. J Cell Biochem 112:98-106
Zhang, Chi; Jia, Pingping; Jia, Yuanyuan et al. (2010) Methionine sulfoxide reductase A (MsrA) protects cultured mouse embryonic stem cells from H2O2-mediated oxidative stress. J Cell Biochem 111:94-103
Zhang, Chi; Jia, Pingping; Huang, Xupei et al. (2009) Myofibril-inducing RNA (MIR) is essential for tropomyosin expression and myofibrillogenesis in axolotl hearts. J Biomed Sci 16:81
Jia, P; Zhang, C; Huang, X P et al. (2008) A novel protein involved in heart development in Ambystoma mexicanum is localized in endoplasmic reticulum. J Biomed Sci 15:789-99
Zhang, C; Pietras, K M; Sferrazza, G F et al. (2007) Molecular and immunohistochemical analyses of cardiac troponin T during cardiac development in the Mexican axolotl, Ambystoma mexicanum. J Cell Biochem 100:1-15
Sferrazza, Gian Franco; Zhang, Chi; Jia, Pingping et al. (2007) Role of myofibril-inducing RNA in cardiac TnT expression in developing Mexican axolotl. Biochem Biophys Res Commun 357:32-7

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