Gata factor Pannier is required to establish competence for heart progenitor formation

Abstract

Inductive signaling is of pivotal importance for developmental patterns to form. In Drosophila, the transfer of TGFβ (Dpp) and Wnt (Wg) signaling information from the ectoderm to the underlying mesoderm induces cardiacspecific differentiation in the presence of Tinman, a mesoderm-specific homeobox transcription factor. We present evidence that the Gata transcription factor, Pannier, and its binding partner U-shaped, also a zincfinger protein, cooperate in the process of heart development. Loss-of-function and germ layer-specific rescue experiments suggest that pannier provides an essential function in the mesoderm for initiation of cardiacspecific expression of tinman and for specification of the heart primordium. u-shaped also promotes heart development, but unlike pannier, only by maintaining tinman expression in the cardiogenic region. By contrast, pan-mesodermal overexpression of pannier ectopically expands tinman expression, whereas overexpression of ushaped inhibits cardiogenesis. Both factors are also required for maintaining dpp expression after germ band retraction in the dorsal ectoderm. Thus, we propose that Pannier mediates as well as maintains the cardiogenic Dpp signal. In support, we find that manipulation of pannier activity in either germ layer affects cardiac specification, suggesting that its function is required in both the mesoderm and the ectoderm.Key words: Drosophila, Heart, Cardiogenesis, Mesoderm, pannier, u-shaped, tinman, dpp 3028 Evans, 1996;Morrisey et al., 1996), where they are thought to regulate cardiac-specific genes (Grepin et al., 1994;Ip et al., 1994; Durocher et al., 1997;Murphy et al., 1997) (reviewed by Molkentin, 2000). Gata4 is already expressed in the early cardiac crescent of the lateral plate mesoderm, and in mice deficient for Gata4, these heart primordia fail to migrate towards the midline where they normally fuse into the primitive heart tube (Molkentin et al., 1997;Kuo et al., 1997). Owing to these ventral closure defects, it has been difficult to discriminate between a direct role for Gata4 in heart formation and an indirect involvement via its function in ventral morphogenesis. Furthermore, Gata4, Gata5 and Gata6 may act in part redundantly, which may further occlude their cardiogenic potential. Consistent with the direct involvement of Gata4 in heart development is the congenital heart disease phenotype observed in individuals heterozygous for deletions of chromosome 8p23.1 region, which includes the GATA4 gene (Pehlivan et al., 1999;Bhatia et al., 1999).In vitro, Gata4 interacts with a wide array of proteins, including the Tinman homolog Nkx2.5, the bHLH protein Hand and the multiple zinc-finger protein Fog2 (Durocher et al., 1997;Sepulveda et al., 1998;Lee et al., 1998;Lu et al., 1999;Sepulveda et al., 2002;Svensson et al., 1999;Tevosian et al., 1999; Dai et al., 2002). Fog2 apparently modulates Gatamediated transcriptional regulation not only as a repressor, but also as an activator, depending on the promoter and on cell type (Lu et al., 1999). Fog2 is co-...

The ATP-sensitive potassium (K ATP ) channel-encoded dSUR gene is required for Drosophila heart function and is regulated by tinman

“…Pnr augments ectopic Tin-induced dSUR expression, as it does with Dmef-2 and dHand (12,14,32), but the Pnr consensus sites in the cardiac enhancer do not seem to be required. Nevertheless, Tin and Pnr act synergistically in activating dSUR both in vitro and in vivo, and forced expression of tin is unable to restore dSUR expression in flies expressing dominant-negative pnr-EnR.…”

“…Whole-mount in situ hybridization was performed as described in ref. 32. Primer sequences for cloning dSUR enhancer fragments, generation of UAS-dSUR-RNAi transgenic flies, and quantitative RT-PCR (Roche) are described in Supporting Materials and Methods.…”

“…Moreover, both dSUR and dHand were strongly activated when tin and pnr were coexpressed (Fig. 3 L and M), suggesting that, like dHand, dSUR activation depends on genetic synergy between Tin and Pnr (14,32).…”

Genetic Interaction of Lobe With Its Modifiers in Dorsoventral Patterning and Growth of the Drosophila Eye

“…, an amorph allele of emc (Brown et al 1995); Bar 1 (B 1 )/FM7, a dominant gain-offunction mutation generated by duplication of the 16A region containing BarH1 and BarH2 genes, referred to as Bar (B); Df(1)Bar , a deficiency in which both B genes are deleted (Higashijima et al 1992); sgg 1 /FM7a Dp(1;2;4) w 1 , a lethal recessive X-ray allele of sgg (Bourouis et al 1990); Df (2L) BSC7/CyO, UAS-pnr D4 (Haenlin et al 1997); y w; FRT82, pnr vx6 / TM6B (Heitzler et al 1996); UAS-pnr ENR , a dominant-negative form of pnr, which contains the repressor domain from Engrailed (amino acids 2-298) (Klinedinst and Bodmer 2003); UAS-ush (Fossett et al 2001); Df (3L) Iro-C DFM3 FRT80/TM6B, a deficiency for ara, caup, and promoter for mirr, UAS-ara (Gomez-Skarmeta et al 1996); UAS-mirr (McNeill et al 1997); y w ey-FLP (Newsome et al 2000); and pnr-GAL4 . All stocks are described in FlyBase (http:/ /flybase.bio.indiana.edu).…”

Patterning of the cardiac outflow region in Drosophila

“…Recent studies of genes involved in heart development (5, 6) have revealed discrete subsets of cardioblasts and pericardial cells. According to the expression domains of heart identity genes, in each abdominal segment three types of cardioblasts can be detected, the anterior tinman (tin)-expressing pair, the adjacent tin͞ladybird (lb)-positive pair, and the most posterior seven up (svp)-expressing pair (5)(6)(7)(8)(9)(10). As for pericardial cells, the combinatory code of tin, lb, even-skipped (eve), and odd-skipped (odd) expression makes it possible to distinguish five distinct subpopulations (6-8, 10, 11).…”