Article

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.

Download full-text

Full-text

Available from: Lionel Christiaen, Aug 30, 2015
0 Followers
 · 
87 Views
  • Source
    • "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). "
    [Show abstract] [Hide abstract]
    ABSTRACT: The study of cis-regulatory DNAs that control developmental gene expression is integral to the modeling of comprehensive genomic regulatory networks for embryogenesis. Ascidian embryos provide a unique opportunity for the analysis of cis-regulatory DNAs with cellular resolution in the context of a simple but typical chordate body plan. Here, we review landmark studies that have laid the foundations for the study of transcriptional enhancers, among other cis-regulatory DNAs, and their roles in ascidian development. The studies using ascidians of the Ciona genus have capitalized on a unique electroporation technique that permits the simultaneous transfection of hundreds of fertilized eggs, which develop rapidly and express transgenes with little mosaicism. Current studies using the ascidian embryo benefit from extensively annotated genomic resources to characterize transcript models in silico. The search for functional noncoding sequences can be guided by bioinformatic analyses combining evolutionary conservation, gene coexpression, and combinations of overrepresented short-sequence motifs. The power of the transient transfection assays has allowed thorough dissection of numerous cis-regulatory modules, which provided insights into the functional constraints that shape enhancer architecture and diversification. Future studies will benefit from pioneering stable transgenic lines and the analysis of chromatin states. Whole genome expression, functional and DNA binding data are being integrated into comprehensive genomic regulatory network models of early ascidian cell specification with a single-cell resolution that is unique among chordate model systems.
    Current Topics in Developmental Biology 01/2012; 98:147-72. DOI:10.1016/B978-0-12-386499-4.00006-9 · 4.21 Impact Factor
  • Source
    • "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. "
    [Show abstract] [Hide abstract]
    ABSTRACT: The heart and other blood pumping organs are close to being universally essential in the animal kingdom. These organs present a large anatomical, morphological, and cellular diversity, which is thought to have arisen by building developmental modules on a conserved core of ancestral heart regulatory units. In this context, studies using the ascidian model system Ciona intestinalis offer a distinctive set of theoretical and experimental advantages, which we herein discuss in details. Development of the heart and related muscles in Ciona has been analyzed with a cellular to subcellular resolution unprecedented in Chordate model systems. Unique derived developmental characters of the cardiogenic mesoderm appear to be shared between Ciona and vertebrates. Notably, accumulating evidence point to an early Chordate origin of the cardiopharyngeal population of mesoderm cells that may have provided the foundation for the emergence of the second heart field in higher vertebrates.
    Current Topics in Developmental Biology 01/2012; 100:107-42. DOI:10.1016/B978-0-12-387786-4.00011-7 · 4.21 Impact Factor
  • Source
    • "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). "
    [Show abstract] [Hide abstract]
    ABSTRACT: For over a century, muscle formation in the ascidian embryo has been representative of 'mosaic' development. The molecular basis of muscle-fate predetermination has been partly elucidated with the discovery of Macho1, a maternal zinc-finger transcription factor necessary and sufficient for primary muscle development, and of its transcriptional intermediaries Tbx6b and Tbx6c. However, the molecular mechanisms by which the maternal information is decoded by cis-regulatory modules (CRMs) associated with muscle transcription factor and structural genes, and the ways by which a seamless transition from maternal to zygotic transcription is ensured, are still mostly unclear. By combining misexpression assays with CRM analyses, we have identified the mechanisms through which Ciona Macho1 (Ci-Macho1) initiates expression of Ci-Tbx6b and Ci-Tbx6c, and we have unveiled the cross-regulatory interactions between the latter transcription factors. Knowledge acquired from the analysis of the Ci-Tbx6b CRM facilitated both the identification of a related CRM in the Ci-Tbx6c locus and the characterization of two CRMs associated with the structural muscle gene fibrillar collagen 1 (CiFCol1). We use these representative examples to reconstruct how compact CRMs orchestrate the muscle developmental program from pre-localized ooplasmic determinants to differentiated larval muscle in ascidian embryos.
    Journal of Cell Science 07/2010; 123(Pt 14):2453-63. DOI:10.1242/jcs.066910 · 5.33 Impact Factor
Show more