Spatio-temporal intersection of Lhx3 and Tbx6 defines the cardiac field through synergistic activation of Mesp

Department of Molecular & Cell Biology, Division of Genetics, Genomics and Development, Center for Integrative Genomics, University of California Berkeley, CA 94720-3200, USA.
Developmental Biology (Impact Factor: 3.64). 03/2009; 328(2):552-60. DOI: 10.1016/j.ydbio.2009.01.033
Source: PubMed

ABSTRACT Mesp encodes a bHLH transcription factor required for specification of the cardiac mesoderm in Ciona embryos. The activities of Macho-1 and beta-catenin, two essential maternal determinants, are required for Mesp expression in the B7.5 blastomeres, which constitute the heart field. The T-box transcription factor Tbx6 functions downstream of Macho-1 as a direct activator of Mesp expression. However, Tbx6 cannot account for the restricted expression of Mesp in the B7.5 lineage since it is expressed throughout the presumptive tail muscles. Here we present evidence that the LIM-homeobox gene Lhx3, a direct target of beta-catenin, is essential for localized Mesp expression. Lhx3 is expressed throughout the presumptive endoderm and B7.5 blastomeres. Thus, the B7.5 blastomeres are the only cells to express sustained levels of the Tbx6 and Lhx3 activators. Like mammalian Lhx3 genes, Ci-Lhx3 encodes two isoforms with distinct N-terminal peptides. The Lhx3a isoform appears to be expressed both maternally and zygotically, while the Lhx3b isoform is exclusively zygotic. Misexpression of Lhx3b is sufficient to induce ectopic Mesp activation in cells expressing Tbx6b. Injection of antisense morpholino oligonucleotides showed that the Lhx3b isoform is required for endogenous Mesp expression. Mutations in the Lhx3 half-site of Tbx6/Lhx3 composite elements strongly reduced the activity of a minimal Mesp enhancer. We discuss the delineation of the heart field by the synergistic action of muscle and gut determinants.

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Available from: Lionel Christiaen, Aug 30, 2015
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    • "Thus, B7.5 blastomeres uniquely activate Mesp transcription because they are the only cells to express the two necessary transactivators Tbx6b/c and Lhx3b in the 64-cell embryo. This synergistic developmental logic is recapitulated on the Mesp cis-regulatory DNA, where partially overlapping Lhx3 and Tbx6 binding sites are required for reporter gene expression in B7.5 blastomeres (Christiaen et al., 2009a). By the tailbud stage, the B7.5 blastomeres give birth to two distinct types of cells: anterior tail muscles (ATMs) and the trunk ventral cells (TVCs), which migrate anteriorly and constitute the heart and atrial siphon muscle progenitors (Davidson and Levine, 2003; Hirano and Nishida, 1997; Satou et al., 2004; Stolfi et al., 2010); Fig. 6.2C). "
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    • "A combination of misexpression and gene knockdown assays showed that a Tbx6b and Lhx3 synergy activated Mesp specifically in B7.5 blastomeres, which are the only cells that coexpress Lhx3 and Tbx6b in the 110-cell stage embryo, when Mesp expression starts (Fig. 4.2; Christiaen et al., 2009a–d). The minimal Mesp enhancer contains putative Lhx3 binding sites that overlap previously identified Tbx6 sites and were required for reporter gene expression in the B7.5 cells (Christiaen et al., 2009a–d). These data indicate that cooperative DNA binding may contribute to the observed synergy between the two necessary trans-activators of Mesp, thus encoding a developmental logic—the restricted overlap of two necessary activators—in the cis-regulatory DNA of an essential heart specification gene. "
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    • "Similarly, in Halocynthia roretzi, another solitary ascidian distantly related to Ciona, Hr-Macho1 has been shown to activate the expression of Hr-Tbx6, as well as that of structural muscle genes, such as Hr-muscle actin (Sawada et al., 2005). In addition to controlling primary muscle formation, Ci-Macho1 acts cooperatively with !-catenin to induce the formation of the heart field by activating Ci-Mesp (Christiaen et al., 2009; Davidson et al., 2005; Satou et al., 2004); the function of Ci-Macho1 in heart specification is also mediated by the Ci-Tbx6-related transcription factors (Christiaen et al., 2009; Davidson et al., 2005). Despite the wealth of information on the gene regulatory network that initiates and sustains muscle development in Ciona and other ascidian embryos (Hudson and Yasuo, 2008; Imai et al., 2006; Meedel et al., 2007), the cis-regulatory mechanisms that integrate maternal and zygotic information along this complex gene cascade, from egg to swimming larva, are largely unexplored, although some common logic has been identified in a limited subset of cisregulatory modules (CRMs) (Erives, 2009). "
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