Claudia Nieva

Capicua DNA-binding sites are general response elements for RTK signaling in Drosophila

RTK/Ras/MAPK signaling pathways play key functions in metazoan development, but how they control expression of downstream genes is not well understood. In Drosophila, it is generally assumed that most transcriptional responses to RTK signal activation depend on binding of Ets-family proteins to specific cis-acting sites in target enhancers. Here, we show that several Drosophila RTK pathways control expression of downstream genes through common octameric elements that are binding sites for the HMG-box factor Capicua, a transcriptional repressor that is downregulated by RTK signaling in different contexts. We show that Torso RTK-dependent regulation of terminal gap gene expression in the early embryo critically depends on Capicua octameric sites, and that binding of Capicua to these sites is essential for recruitment of the Groucho co-repressor to the huckebein enhancer in vivo. We then show that subsequent activation of the EGFR RTK pathway in the neuroectodermal region of the embryo controls dorsal-ventral gene expression by downregulating the Capicua protein, and that this control also depends on Capicua octameric motifs. Thus, a similar mechanism of RTK regulation operates during subdivision of the anterior-posterior and dorsal-ventral embryonic axes. We also find that identical DNA octamers mediate Capicua-dependent regulation of another EGFR target in the developing wing. Remarkably, a simple combination of activator-binding sites and Capicua motifs is sufficient to establish complex patterns of gene expression in response to both Torso and EGFR activation in different tissues. We conclude that Capicua octamers are general response elements for RTK signaling in Drosophila.

Isolation and Functional Characterisation of Two New bZIP Maize Regulators of the ABA Responsive Gene rab28

The plant hormone abscisic acid regulates gene expression in response to growth stimuli and abiotic stress. Previous studies have implicated members of the bZIP family of transcription factors as mediators of abscisic acid dependent gene expression through the ABRE cis-element. Here, we identify two new maize bZIP transcription factors, EmBP-2 and ZmBZ-1 related to EmBP-1 and OsBZ-8 families. They are differentially expressed during embryo development; EmBP-2 is constitutive, whereas ZmBZ-1 is abscisic acid-inducible and accumulates during late embryogenesis. Both factors are nuclear proteins that bind to ABREs and activate transcription of the abscisic acid-inducible gene rab28 from maize. EmBP-2 and ZmBZ-1 are phosphorylated by protein kinase CK2 and phosphorylation alters their DNA binding properties. Our data suggest that EmBP-2 and ZmBZ-1 are involved in the expression of abscisic acid inducible genes such as rab28 and their activity is modulated by ABA and by phosphorylation.

Nuclear receptor BgFTZ‐F1 regulates molting and the timing of ecdysteroid production during nymphal development in the hemimetabolous insect Blattella germanica

Postembryonic development of holometabolous and hemimetabolous insects occurs through successive molts triggered by 20‐hydroxyecdysone (20E). The molecular action of 20E has been extensively studied in holometabolous insects, but data on hemimetabolous are scarce. We have demonstrated that during the nymphal development of the hemimetabolous insect Blattella germanica, 20E binds to the heterodimeric receptor formed by the nuclear receptors BgEcR‐A and BgRXR activating a cascade of gene expression, including the nuclear receptors BgE75 and BgHR3. Herein, we report the characterization of BgFTZ‐F1, another nuclear hormone receptor involved in 20E action. BgFTZ‐F1 is activated at the end of each instar, and RNAi has demonstrated that BgHR3 is needed for BgFTZ‐F1 activation, and that BgFTZ‐F1 has critical functions of during the last nymphal instar. Nymphs with silenced BgFTZ‐F1 cannot ecdyse, arrest development, and show structures of ectodermal origin duplicated. BgFTZ‐F1 also controls the timing of the ecdysteroid molting pulse. Developmental Dynamics 237:3179–3191, 2008.

The hormonal pathway controlling cell death during metamorphosis in a hemimetabolous insect

Metamorphosis in holometabolous insects is mainly based on the destruction of larval tissues. Intensive research in Drosophila melanogaster, a model of holometabolan metamorphosis, has shown that the steroid hormone 20-hydroxyecdysone (20E) signals cell death of larval tissues during metamorphosis. However, D. melanogaster shows a highly derived type of development and the mechanisms regulating apoptosis may not be representative in the insect class context. Unfortunately, no functional studies have been carried out to address whether the mechanisms controlling cell death are present in more basal hemimetabolous species. To address this, we have analyzed the apoptosis of the prothoracic gland of the cockroach Blattella germanica, which undergoes stage-specific degeneration just after the imaginal molt. Here, we first show that B. germanica has two inhibitor of apoptosis (IAP) proteins and that one of them, BgIAP1, is continuously required to ensure tissue viability, including that of the prothoracic gland, during nymphal development. Moreover, we demonstrate that the degeneration of the prothoracic gland is controlled by a complex 20E-triggered hierarchy of nuclear receptors converging in the strong activation of the death-inducer Fushi tarazu-factor 1 (BgFTZ-F1) during the nymphal-adult transition. Finally, we have also shown that prothoracic gland degeneration is effectively prevented by the presence of juvenile hormone (JH). Given the relevance of cell death in the metamorphic process, the characterization of the molecular mechanisms regulating apoptosis in hemimetabolous insects would allow to help elucidate how metamorphosis has evolved from less to more derived insect species.

Origins of context-dependent gene repression by capicua.

Receptor Tyrosine Kinase (RTK) signaling pathways induce multiple biological responses, often by regulating the expression of downstream genes. The HMG-box protein Capicua (Cic) is a transcriptional repressor that is downregulated in response to RTK signaling, thereby enabling RTK-dependent induction of Cic targets. In both Drosophila and mammals, Cic is expressed as two isoforms, long (Cic-L) and short (Cic-S), whose functional significance and mechanism of action are not well understood. Here we show that Drosophila Cic relies on the Groucho (Gro) corepressor during its function in the early embryo, but not during other stages of development. This Gro-dependent mechanism requires a short peptide motif, unique to Cic-S and designated N2, which is distinct from other previously defined Gro-interacting motifs and functions as an autonomous, transferable repressor element. Unexpectedly, our data indicate that the N2 motif is an evolutionary innovation that originated within dipteran insects, as the Cic-S isoform evolved from an ancestral Cic-L-type form. Accordingly, the Cic-L isoform lacking the N2 motif is completely inactive in early Drosophila embryos, indicating that the N2 motif endowed Cic-S with a novel Gro-dependent activity that is obligatory at this stage. We suggest that Cic-S and Gro coregulatory functions have facilitated the evolution of the complex transcriptional network regulated by Torso RTK signaling in modern fly embryos. Notably, our results also imply that mammalian Cic proteins are unlikely to act via Gro and that their Cic-S isoform must have evolved independently of fly Cic-S. Thus, Cic proteins employ distinct repressor mechanisms that are associated with discrete structural changes in the evolutionary history of this protein family.

Juvenile hormone biosynthesis in adult Blattella germanica requires nuclear receptors Seven-up and FTZ-F1

In insects, the transition from juvenile development to the adult stage is controlled by juvenile hormone (JH) synthesized from the corpora allata (CA) glands. Whereas a JH-free period during the last juvenile instar triggers metamorphosis and the end of the growth period, the reappearance of this hormone after the imaginal molt marks the onset of reproductive adulthood. Despite the importance of such transition, the regulatory mechanism that controls it remains mostly unknown. Here, using the hemimetabolous insect Blattella germanica, we show that nuclear hormone receptors Seven-up-B (BgSvp-B) and Fushi tarazu-factor 1 (BgFTZ-F1) have essential roles in the tissue- and stage-specific activation of adult CA JH-biosynthetic activity. Both factors are highly expressed in adult CA cells. Moreover, RNAi-knockdown of either BgSvp-B or BgFTZ-F1 results in adult animals with a complete block in two critical JH-dependent reproductive processes, vitellogenesis and oogenesis. We show that this reproductive blockage is the result of a dramatic impairment of JH biosynthesis, due to the CA-specific reduction in the expression of two key JH biosynthetic enzymes, 3-hydroxy-3-methylglutaryl coenzyme A synthase-1 (BgHMG-S1) and HMG-reductase (BgHMG-R). Our findings provide insights into the regulatory mechanisms underlying the specific changes in the CA gland necessary for the proper transition to adulthood.

PLANT AP2/EREBP AND bZIP TRANSCRIPTION FACTORS: STRUCTURE AND FUNCTION

Regulation of gene expression by transcription factors is a central mechanism utilized among organisms and relies on interactions between sequence-specific DNA-binding proteins with cis-elements located in the promoter and enhancer regions of the corresponding genes. In plants, a large number of transcription factors has been identified the past years, with some of them exhibiting structural features unique to plant transcription factors. In this review we summarize the current knowledge on the AP2/EREBP and bZIP families of plant transcription factors stressing out putative ways of their mode of regulation and function.