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The Drosophila Transcription Factors Tinman and Pannier Activate and Collaborate with Myocyte Enhancer Factor-2 to Promote Heart Cell Fate.

Lovato TL, Sensibaugh CA, Swingle KL, Martinez MM, Cripps RM - PLoS ONE (2015)

Bottom Line: We found that mesodermal over-expression of Tinman and Pannier resulted in approximately 20% of embryos with ectopic Hand and Sulphonylurea receptor (Sur) expression.By adding MEF2 alongside Tinman and Pannier, a dramatic expansion in the expression of Hand and Sur was observed in almost all embryos analyzed.Two additional cardiac markers were also expanded in their expression.

View Article: PubMed Central - PubMed

Affiliation: Department of Biology, University of New Mexico, Albuquerque, NM 87131-1091, United States of America.

ABSTRACT
Expression of the MADS domain transcription factor Myocyte Enhancer Factor 2 (MEF2) is regulated by numerous and overlapping enhancers which tightly control its transcription in the mesoderm. To understand how Mef2 expression is controlled in the heart, we identified a late stage Mef2 cardiac enhancer that is active in all heart cells beginning at stage 14 of embryonic development. This enhancer is regulated by the NK-homeodomain transcription factor Tinman, and the GATA transcription factor Pannier through both direct and indirect interactions with the enhancer. Since Tinman, Pannier and MEF2 are evolutionarily conserved from Drosophila to vertebrates, and since their vertebrate homologs can convert mouse fibroblast cells to cardiomyocytes in different activator cocktails, we tested whether over-expression of these three factors in vivo could ectopically activate known cardiac marker genes. We found that mesodermal over-expression of Tinman and Pannier resulted in approximately 20% of embryos with ectopic Hand and Sulphonylurea receptor (Sur) expression. By adding MEF2 alongside Tinman and Pannier, a dramatic expansion in the expression of Hand and Sur was observed in almost all embryos analyzed. Two additional cardiac markers were also expanded in their expression. Our results demonstrate the ability to initiate ectopic cardiac fate in vivo by the combination of only three members of the conserved Drosophila cardiac transcription network, and provide an opportunity for this genetic model system to be used to dissect the mechanisms of cardiac specification.

No MeSH data available.


Related in: MedlinePlus

Expression of tinman, pannier and Mef2 in the ectoderm results in an expansion of cardiac gene expression.(A,B) 69B>tin+pnr+Mef2 embryos at stage 14, stained for Hand expression (A) or Sur expression (B). (A) ~ 50% of embryos had expanded Hand expression (arrowhead) with the rest demonstrating normal expression in the heart (inset, arrow). (B) ~70% of embryos had normal Sur expression in the heart (arrow), with a subset showing expanded Sur expression in the nervous system (inset, arrowhead). Bar, 100μm. (C) Quantification of effects of over-expression of cardiac transcription factors. (D,E) Antibody stain against Fasciclin III. (D) shows normal FasIII accumulation in a stage 9 control embryo. (E) shows similar levels of accumulation in a 69B>tin+pnr+Mef2 embryo.
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pone.0132965.g007: Expression of tinman, pannier and Mef2 in the ectoderm results in an expansion of cardiac gene expression.(A,B) 69B>tin+pnr+Mef2 embryos at stage 14, stained for Hand expression (A) or Sur expression (B). (A) ~ 50% of embryos had expanded Hand expression (arrowhead) with the rest demonstrating normal expression in the heart (inset, arrow). (B) ~70% of embryos had normal Sur expression in the heart (arrow), with a subset showing expanded Sur expression in the nervous system (inset, arrowhead). Bar, 100μm. (C) Quantification of effects of over-expression of cardiac transcription factors. (D,E) Antibody stain against Fasciclin III. (D) shows normal FasIII accumulation in a stage 9 control embryo. (E) shows similar levels of accumulation in a 69B>tin+pnr+Mef2 embryo.

Mentions: To determine if Tin, Pnr and MEF2 can potentiate the cardiac phenotype outside of the mesoderm, we tested whether they could activate the cardiac program in the ectoderm, using the 69B-gal4 driver line. We found that, when we used Hand as a marker of heart fate, a little over 50% of the embryos stained had ectopic stain; and when Sur was used as a cardiac marker, 30% of the embryos had ectopic expression in ectodermal tissues (Fig 7A and 7B). Sur expression was only narrowly expanded to what appeared to be malformed salivary glands, which arise from ectodermal cells. In addition, there was no expansion of FasIII accumulation in stage 12 embryos, indicating no expansion of visceral mesoderm fate in these embryos (Fig 7D and 7E). These results suggested that conversion to a cardiac fate requires either some threshold level of activation by the converting factors that may or may not be met when utilizing an ectodermal driver; or, there are additional mesodermal factors with which Tin, Pnr and MEF2 collaborate to activate the myogenic program.


The Drosophila Transcription Factors Tinman and Pannier Activate and Collaborate with Myocyte Enhancer Factor-2 to Promote Heart Cell Fate.

Lovato TL, Sensibaugh CA, Swingle KL, Martinez MM, Cripps RM - PLoS ONE (2015)

Expression of tinman, pannier and Mef2 in the ectoderm results in an expansion of cardiac gene expression.(A,B) 69B>tin+pnr+Mef2 embryos at stage 14, stained for Hand expression (A) or Sur expression (B). (A) ~ 50% of embryos had expanded Hand expression (arrowhead) with the rest demonstrating normal expression in the heart (inset, arrow). (B) ~70% of embryos had normal Sur expression in the heart (arrow), with a subset showing expanded Sur expression in the nervous system (inset, arrowhead). Bar, 100μm. (C) Quantification of effects of over-expression of cardiac transcription factors. (D,E) Antibody stain against Fasciclin III. (D) shows normal FasIII accumulation in a stage 9 control embryo. (E) shows similar levels of accumulation in a 69B>tin+pnr+Mef2 embryo.
© Copyright Policy
Related In: Results  -  Collection

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Show All Figures
getmorefigures.php?uid=PMC4520567&req=5

pone.0132965.g007: Expression of tinman, pannier and Mef2 in the ectoderm results in an expansion of cardiac gene expression.(A,B) 69B>tin+pnr+Mef2 embryos at stage 14, stained for Hand expression (A) or Sur expression (B). (A) ~ 50% of embryos had expanded Hand expression (arrowhead) with the rest demonstrating normal expression in the heart (inset, arrow). (B) ~70% of embryos had normal Sur expression in the heart (arrow), with a subset showing expanded Sur expression in the nervous system (inset, arrowhead). Bar, 100μm. (C) Quantification of effects of over-expression of cardiac transcription factors. (D,E) Antibody stain against Fasciclin III. (D) shows normal FasIII accumulation in a stage 9 control embryo. (E) shows similar levels of accumulation in a 69B>tin+pnr+Mef2 embryo.
Mentions: To determine if Tin, Pnr and MEF2 can potentiate the cardiac phenotype outside of the mesoderm, we tested whether they could activate the cardiac program in the ectoderm, using the 69B-gal4 driver line. We found that, when we used Hand as a marker of heart fate, a little over 50% of the embryos stained had ectopic stain; and when Sur was used as a cardiac marker, 30% of the embryos had ectopic expression in ectodermal tissues (Fig 7A and 7B). Sur expression was only narrowly expanded to what appeared to be malformed salivary glands, which arise from ectodermal cells. In addition, there was no expansion of FasIII accumulation in stage 12 embryos, indicating no expansion of visceral mesoderm fate in these embryos (Fig 7D and 7E). These results suggested that conversion to a cardiac fate requires either some threshold level of activation by the converting factors that may or may not be met when utilizing an ectodermal driver; or, there are additional mesodermal factors with which Tin, Pnr and MEF2 collaborate to activate the myogenic program.

Bottom Line: We found that mesodermal over-expression of Tinman and Pannier resulted in approximately 20% of embryos with ectopic Hand and Sulphonylurea receptor (Sur) expression.By adding MEF2 alongside Tinman and Pannier, a dramatic expansion in the expression of Hand and Sur was observed in almost all embryos analyzed.Two additional cardiac markers were also expanded in their expression.

View Article: PubMed Central - PubMed

Affiliation: Department of Biology, University of New Mexico, Albuquerque, NM 87131-1091, United States of America.

ABSTRACT
Expression of the MADS domain transcription factor Myocyte Enhancer Factor 2 (MEF2) is regulated by numerous and overlapping enhancers which tightly control its transcription in the mesoderm. To understand how Mef2 expression is controlled in the heart, we identified a late stage Mef2 cardiac enhancer that is active in all heart cells beginning at stage 14 of embryonic development. This enhancer is regulated by the NK-homeodomain transcription factor Tinman, and the GATA transcription factor Pannier through both direct and indirect interactions with the enhancer. Since Tinman, Pannier and MEF2 are evolutionarily conserved from Drosophila to vertebrates, and since their vertebrate homologs can convert mouse fibroblast cells to cardiomyocytes in different activator cocktails, we tested whether over-expression of these three factors in vivo could ectopically activate known cardiac marker genes. We found that mesodermal over-expression of Tinman and Pannier resulted in approximately 20% of embryos with ectopic Hand and Sulphonylurea receptor (Sur) expression. By adding MEF2 alongside Tinman and Pannier, a dramatic expansion in the expression of Hand and Sur was observed in almost all embryos analyzed. Two additional cardiac markers were also expanded in their expression. Our results demonstrate the ability to initiate ectopic cardiac fate in vivo by the combination of only three members of the conserved Drosophila cardiac transcription network, and provide an opportunity for this genetic model system to be used to dissect the mechanisms of cardiac specification.

No MeSH data available.


Related in: MedlinePlus