The mammalian TEF and the Drosophila scalloped genes belong to a conserved family of transcriptional factors that possesses a TEA/ATTS DNA-binding domain. Transcriptional activation by these proteins likely requires interactions with specific coactivators. In Drosophila, Scalloped (Sd) interacts with Vestigial (Vg) to form a complex that binds DNA through the Sd TEA/ATTS domain. The Sd-Vg heterodimer is a key regulator of wing development; this heterodimer directly controls several target genes and is able to induce wing outgrowth when ectopically expressed. Vg contains two distinct transcriptional activation domains, suggesting that the function of Vg is to mediate transcriptional activation by Sd. By expressing a chimeric GAL4-Sd protein in Drosophila, it is found that the transcriptional activity of the Vg-Sd heterodimer is negatively regulated at the AP and DV boundary of the wing disc. A novel human protein, TONDU, is identified that contains a short domain homologous to the domain of Vg required for interaction with Sd. TONDU specifically interacts with a domain conserved in all the mammalian TEF factors. Expression of TDU in Drosophila by means of the UAS-GAL4 system shows that this human protein can substitute for Vg in wing formation. It is proposed that TDU is a specific coactivator for the mammalian TEFs (Vaudin, 1999).
The Sd/Vg dimer is active at the DV boundary at the end of the second larval instar, but is strongly downregulated at the DV and AP boundaries in late imaginal disc development. Importantly, because these experiments were done without ectopic gene expression, they probably accurately reflect the wild-type situation for vg and sd. It is proposed that the function of the Sd/Vg heterodimer is not solely to restrict the expression of target genes to the wing pouch, but that it also participates in setting up the pattern of gene expression within the wing pouch. The modulation of Vg/Sd activity could also contribute to growth control during wing development. Indeed, in the wing disc, a zone of non-proliferating cells (ZNC), coinciding with the DV boundary of the wing pouch, is formed during the third instar. This ZNC is established through the coordinate action of Notch and Wg and is required for a correct specification of the wing margin. Several lines of evidence point to a role for vg in the control of cell proliferation: clones with a loss of vg function fail to proliferate, while ectopic vg expression induces cell proliferation. Recently, it has been shown that cell proliferation of wing pouch cells controlled by the EGF receptor pathway is mediated by regulation of vg expression (Nagaraj, 1999). Since vg induces cell proliferation, a negative modulation in Vg/Sd activity at the DV boundary could be a prerequisite to the formation of the ZNC (Vaudin, 1999).
These results highlight the advantage of using a transcription factor complex combining a DNA-binding molecule (Sd) and an activator molecule (Vg) in order to dynamically control the activity of several cognate enhancers that have to be precisely and dynamically expressed during wing growth and determination. The activity of such transcription factor complexes can be modulated by changing the expression level of either component. In the case of Sd and Vg, an increase in Sd or a reduction in Vg may lead to diminished activation, because Sd alone can bind to its cognate sites and act as a dominant negative inhibitor of the Sd/Vg complex. Such a situation increases the combinatorial possibilities for the fine tuning of transcriptional regulation (Vaudin, 1999).
Sd belongs to an evolutionary conserved family of transcriptional factors. In mammals, the functions of these different TEFs are not completely understood, but they are probably involved in the transcriptional control of cardiac, skeletal muscle and placental genes. Indeed, it has been shown that the TEFs bind to the M-CAT element found in the regulatory region of several cardiac and skeletal muscle genes and the placenta-specific chorionic somatomamotrophin gene (hCS). Furthermore, disruption of the TEF-1 gene (Tead1) in mouse is associated with heart defects showing that TEF-1 is likely to be involved in the regulation of specific cardiac genes. Like Sd, TEF function may depend on interaction with coactivators that are expressed in a specific pattern to activate transcription. Indeed transactivation by TEF-1 is cell-type dependent and could act as a transcriptional repressor in particular cell types. In addition, two factors, NEF-1 and NEF-2, negatively regulate transcriptional activation by TEF-1 in vitro, but none of these factors have been cloned. The results showing that TDU interacts with the TEF factors suggest that TDU is a good candidate coactivator. TDU is expressed in lung, kidney and placenta, and also probably in fetal heart since ESTs for TDU were originally identified in these tissues. Heart, skeletal muscle, lung, placenta and kidney are enriched in transcripts for various TEFs. It is therefore possible that TDU acts as a transcriptional coactivator for the TEFs in these tissues. allowing the activation of specific target genes (Vaudin, 1999).
Expression of many skeletal muscle-specific genes depends on TEF-1 (transcription enhancer factor-1) and MEF2 transcription factors. In Drosophila, the TEF-1 homolog Scalloped interacts with the cofactor Vestigial to drive differentiation of the wing and indirect flight muscles. Three mammalian vestigial-like genes, Vgl-1, Vgl-2, and Vgl-3, have been identified that share homology in a TEF-1 interaction domain. Vgl-1 and Vgl-3 transcripts are enriched in the placenta, whereas Vgl-2 is expressed in the differentiating somites and branchial arches during embryogenesis and is skeletal muscle-specific in the adult. During muscle differentiation, Vgl-2 mRNA levels increase and Vgl-2 protein translocates from the cytoplasm to the nucleus. In situ hybridization revealed co-expression of Vgl-2 with myogenin in the differentiating muscle of embryonic myotomes but not in newly formed somites prior to muscle differentiation. Like Vgl-1, Vgl-2 interacts with TEF-1. In addition, Vgl-2 interacts with MEF2 in a mammalian two-hybrid assay and Vgl-2 selectively binds to MEF2 in vitro. Co-expression of Vgl-2 with MEF2 markedly co-activates an MEF2-dependent promoter through its MEF2 element. Overexpression of Vgl-2 in MyoD-transfected 10T(1/2) cells markedly increases myosin heavy chain expression, a marker of terminal muscle differentiation. These results identify Vgl-2 as an important new component of the myogenic program (Maeda, 2002).
The co-factor Vestigial-like 2 (Vgl-2), in association with the Scalloped/Tef/Tead transcription factors, has been identified as a component of the myogenic program in the C2C12 cell line. In order to understand Vgl-2 function in embryonic muscle formation, Vgl-2 expression and regulation were analyzed during chick embryonic development. Vgl-2 expression was associated with all known sites of skeletal muscle formation, including those in the head, trunk and limb. Vgl-2 was expressed after the myogenic factor MyoD, regardless of the site of myogenesis. Analysis of Vgl-2 regulation by Notch signalling showed that Vgl-2 expression was down-regulated by Delta1-activated Notch, similarly to the muscle differentiation genes MyoD, Myogenin,Desmin, and Mef2c, while the expression of the muscle progenitor markers such as Myf5, Six1 and FgfR4 was not modified. Moreover, it was established that the Myogenic Regulatory Factors (MRFs) associated with skeletal muscle differentiation (MyoD, Myogenin and Mrf4) were sufficient to activate Vgl-2 expression, while Myf5 was not able to do so. The Vgl-2 endogenous expression, the similar regulation of Vgl-2 and that of MyoD and Myogenin by Notch signalling, and the positive regulation of Vgl-2 by these MRFs suggest that Vgl-2 acts downstream of MyoD activation and is associated with the differentiation step in embryonic skeletal myogenesis (Bonnet, 2010).
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