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Tbx2 and Tbx3 regulate the dynamics of cell proliferation during heart remodeling.

Ribeiro I, Kawakami Y, Büscher D, Raya A, Rodríguez-León J, Morita M, Rodríguez Esteban C, Izpisúa Belmonte JC - PLoS ONE (2007)

Bottom Line: Cardiomyocytes in the future chamber myocardium acquire different cellular and physiological characteristics through activation of a chamber-specific genetic program, which is in part mediated by T-box genes.We characterize two new zebrafish T-box transcription factors, tbx3b and tbx2a, and analyze their role during the development of the atrioventricular canal.Loss- and gain-of-function analyses demonstrate that tbx3b and tbx2a are necessary to repress the chamber-genetic program in the non-chamber myocardium.

View Article: PubMed Central - PubMed

Affiliation: Gene Expression Laboratory, The Salk Institute for Biological Studies, La Jolla, California, United States of America.

ABSTRACT

Background: The heart forms from a linear tube that is subject to complex remodeling during embryonic development. Hallmarks of this remodeling are the looping of the heart tube and the regionalization into chamber and non-chamber myocardium. Cardiomyocytes in the future chamber myocardium acquire different cellular and physiological characteristics through activation of a chamber-specific genetic program, which is in part mediated by T-box genes.

Methodology/principal finding: We characterize two new zebrafish T-box transcription factors, tbx3b and tbx2a, and analyze their role during the development of the atrioventricular canal. Loss- and gain-of-function analyses demonstrate that tbx3b and tbx2a are necessary to repress the chamber-genetic program in the non-chamber myocardium. We also show that tbx3b and tbx2a are required to control cell proliferation in the atrioventricular canal and that misregulation of cell proliferation in the heart tube influences looping. Furthermore, we characterize the heart phenotype of a novel Tbx3 mutation in mice and show that both the control of cell proliferation and the repression of chamber-specific genetic program in the non-chamber myocardium are conserved roles of Tbx3 in this species.

Conclusions/significance: Taken together, our results uncover an evolutionarily conserved role of Tbx2/3 transcription factors during remodeling of the heart myocardium and highlight the importance of controlling cell proliferation as a driving force of morphogenesis.

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Aberrant morphology of the AVC in the absence of zebrafish tbx3b and tbx2a.(A–D) Whole mount RNA in situ hybridization of zebrafish tbx3b and tbx2a expression; ventral views show the heart in its maximum extension from the anterior to the posterior pole. Zebrafish tbx3b is expressed throughout the extent of the heart tube at 31 hpf (A) and becomes restricted to the AVC at 42 hpf (B). Zebrafish tbx2a is expressed al low levels throughout the extent of the linear heart tube at 31 hpf (C). At 42 hpf tbx2a transcripts are present in the AVC at high levels (D). Black arrow points to the AVC. (E–M) Ventral views of the heart of a mlc2a::GFP transgenic line that expresses GFP in the myocardium, at 48 hpf (E–H) and 72 hpf (I–M). (E) In wild type 48 hpf embryos, the atrium has moved upward and is positioned at the same anterior-posterior level as the ventricle; (I) later the atrium becomes localized dorsal to the ventricle at 72 hpf. (F, J) Injection of 2.5 ng of tbx3b morpholino into one-cell stage embryos results in delayed heart looping and abnormal AVC. (G, L) Injection of 10 ng of tbx2a MO results in a similar delay in heart looping and an enlargement of the AVC. (H, M) Injection of both tbx3b and tbx2a MOs, at 1.75 and 5 ng respectively, results in the failure to form the AVC constriction and absence of looping. Red arrow indicates the AVC. a, atrium; h, heart; v, ventricle.
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pone-0000398-g001: Aberrant morphology of the AVC in the absence of zebrafish tbx3b and tbx2a.(A–D) Whole mount RNA in situ hybridization of zebrafish tbx3b and tbx2a expression; ventral views show the heart in its maximum extension from the anterior to the posterior pole. Zebrafish tbx3b is expressed throughout the extent of the heart tube at 31 hpf (A) and becomes restricted to the AVC at 42 hpf (B). Zebrafish tbx2a is expressed al low levels throughout the extent of the linear heart tube at 31 hpf (C). At 42 hpf tbx2a transcripts are present in the AVC at high levels (D). Black arrow points to the AVC. (E–M) Ventral views of the heart of a mlc2a::GFP transgenic line that expresses GFP in the myocardium, at 48 hpf (E–H) and 72 hpf (I–M). (E) In wild type 48 hpf embryos, the atrium has moved upward and is positioned at the same anterior-posterior level as the ventricle; (I) later the atrium becomes localized dorsal to the ventricle at 72 hpf. (F, J) Injection of 2.5 ng of tbx3b morpholino into one-cell stage embryos results in delayed heart looping and abnormal AVC. (G, L) Injection of 10 ng of tbx2a MO results in a similar delay in heart looping and an enlargement of the AVC. (H, M) Injection of both tbx3b and tbx2a MOs, at 1.75 and 5 ng respectively, results in the failure to form the AVC constriction and absence of looping. Red arrow indicates the AVC. a, atrium; h, heart; v, ventricle.

Mentions: We isolated two novel Tbx genes in zebrafish, which belong to the Tbx2 subfamily. Based on sequence similarity and expression patterns we named them tbx3b and tbx2a (Fig. S1). tbx3b encodes a putative protein of 694 amino acid residues with overall similarity to human TBX3 of 67.8% and to mouse Tbx3 protein of 67.3%. tbx2a encodes a putative protein of 687 amino acid residues with 61.8% similarity to mouse Tbx2, 63.1% to human TBX2 and 78.2% to zebrafish tbx2b. During heart development, tbx3b is first expressed throughout the extent of the heart tube and becomes restricted to the AVC and OFT at 33 hpf (Fig. 1A,B). Zebrafish tbx2a is first weakly expressed throughout the extent of the heart tube becoming later restricted to the AVC (Fig. 1C,D). Thus, the domain of expression of tbx3b and tbx2a in the heart is similar to that of the mouse counterparts [10], [22].


Tbx2 and Tbx3 regulate the dynamics of cell proliferation during heart remodeling.

Ribeiro I, Kawakami Y, Büscher D, Raya A, Rodríguez-León J, Morita M, Rodríguez Esteban C, Izpisúa Belmonte JC - PLoS ONE (2007)

Aberrant morphology of the AVC in the absence of zebrafish tbx3b and tbx2a.(A–D) Whole mount RNA in situ hybridization of zebrafish tbx3b and tbx2a expression; ventral views show the heart in its maximum extension from the anterior to the posterior pole. Zebrafish tbx3b is expressed throughout the extent of the heart tube at 31 hpf (A) and becomes restricted to the AVC at 42 hpf (B). Zebrafish tbx2a is expressed al low levels throughout the extent of the linear heart tube at 31 hpf (C). At 42 hpf tbx2a transcripts are present in the AVC at high levels (D). Black arrow points to the AVC. (E–M) Ventral views of the heart of a mlc2a::GFP transgenic line that expresses GFP in the myocardium, at 48 hpf (E–H) and 72 hpf (I–M). (E) In wild type 48 hpf embryos, the atrium has moved upward and is positioned at the same anterior-posterior level as the ventricle; (I) later the atrium becomes localized dorsal to the ventricle at 72 hpf. (F, J) Injection of 2.5 ng of tbx3b morpholino into one-cell stage embryos results in delayed heart looping and abnormal AVC. (G, L) Injection of 10 ng of tbx2a MO results in a similar delay in heart looping and an enlargement of the AVC. (H, M) Injection of both tbx3b and tbx2a MOs, at 1.75 and 5 ng respectively, results in the failure to form the AVC constriction and absence of looping. Red arrow indicates the AVC. a, atrium; h, heart; v, ventricle.
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Related In: Results  -  Collection

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getmorefigures.php?uid=PMC1851989&req=5

pone-0000398-g001: Aberrant morphology of the AVC in the absence of zebrafish tbx3b and tbx2a.(A–D) Whole mount RNA in situ hybridization of zebrafish tbx3b and tbx2a expression; ventral views show the heart in its maximum extension from the anterior to the posterior pole. Zebrafish tbx3b is expressed throughout the extent of the heart tube at 31 hpf (A) and becomes restricted to the AVC at 42 hpf (B). Zebrafish tbx2a is expressed al low levels throughout the extent of the linear heart tube at 31 hpf (C). At 42 hpf tbx2a transcripts are present in the AVC at high levels (D). Black arrow points to the AVC. (E–M) Ventral views of the heart of a mlc2a::GFP transgenic line that expresses GFP in the myocardium, at 48 hpf (E–H) and 72 hpf (I–M). (E) In wild type 48 hpf embryos, the atrium has moved upward and is positioned at the same anterior-posterior level as the ventricle; (I) later the atrium becomes localized dorsal to the ventricle at 72 hpf. (F, J) Injection of 2.5 ng of tbx3b morpholino into one-cell stage embryos results in delayed heart looping and abnormal AVC. (G, L) Injection of 10 ng of tbx2a MO results in a similar delay in heart looping and an enlargement of the AVC. (H, M) Injection of both tbx3b and tbx2a MOs, at 1.75 and 5 ng respectively, results in the failure to form the AVC constriction and absence of looping. Red arrow indicates the AVC. a, atrium; h, heart; v, ventricle.
Mentions: We isolated two novel Tbx genes in zebrafish, which belong to the Tbx2 subfamily. Based on sequence similarity and expression patterns we named them tbx3b and tbx2a (Fig. S1). tbx3b encodes a putative protein of 694 amino acid residues with overall similarity to human TBX3 of 67.8% and to mouse Tbx3 protein of 67.3%. tbx2a encodes a putative protein of 687 amino acid residues with 61.8% similarity to mouse Tbx2, 63.1% to human TBX2 and 78.2% to zebrafish tbx2b. During heart development, tbx3b is first expressed throughout the extent of the heart tube and becomes restricted to the AVC and OFT at 33 hpf (Fig. 1A,B). Zebrafish tbx2a is first weakly expressed throughout the extent of the heart tube becoming later restricted to the AVC (Fig. 1C,D). Thus, the domain of expression of tbx3b and tbx2a in the heart is similar to that of the mouse counterparts [10], [22].

Bottom Line: Cardiomyocytes in the future chamber myocardium acquire different cellular and physiological characteristics through activation of a chamber-specific genetic program, which is in part mediated by T-box genes.We characterize two new zebrafish T-box transcription factors, tbx3b and tbx2a, and analyze their role during the development of the atrioventricular canal.Loss- and gain-of-function analyses demonstrate that tbx3b and tbx2a are necessary to repress the chamber-genetic program in the non-chamber myocardium.

View Article: PubMed Central - PubMed

Affiliation: Gene Expression Laboratory, The Salk Institute for Biological Studies, La Jolla, California, United States of America.

ABSTRACT

Background: The heart forms from a linear tube that is subject to complex remodeling during embryonic development. Hallmarks of this remodeling are the looping of the heart tube and the regionalization into chamber and non-chamber myocardium. Cardiomyocytes in the future chamber myocardium acquire different cellular and physiological characteristics through activation of a chamber-specific genetic program, which is in part mediated by T-box genes.

Methodology/principal finding: We characterize two new zebrafish T-box transcription factors, tbx3b and tbx2a, and analyze their role during the development of the atrioventricular canal. Loss- and gain-of-function analyses demonstrate that tbx3b and tbx2a are necessary to repress the chamber-genetic program in the non-chamber myocardium. We also show that tbx3b and tbx2a are required to control cell proliferation in the atrioventricular canal and that misregulation of cell proliferation in the heart tube influences looping. Furthermore, we characterize the heart phenotype of a novel Tbx3 mutation in mice and show that both the control of cell proliferation and the repression of chamber-specific genetic program in the non-chamber myocardium are conserved roles of Tbx3 in this species.

Conclusions/significance: Taken together, our results uncover an evolutionarily conserved role of Tbx2/3 transcription factors during remodeling of the heart myocardium and highlight the importance of controlling cell proliferation as a driving force of morphogenesis.

Show MeSH
Related in: MedlinePlus