Apoptotic cells can induce compensatory cell proliferation through the JNK and the Wingless signaling pathways

In many metazoans, damaged and potentially dangerous cells are rapidly eliminated by apoptosis. In Drosophila, this is often compensated for by extraproliferation of neighboring cells, which allows the organism to tolerate considerable cell death without compromising development and body size. Despite its importance, the mechanistic basis of such compensatory proliferation remains poorly understood. Here, we show that apoptotic cells express the secretory factors wingless (wg) and decapentaplegic (dpp). When cells undergoing apoptosis were kept alive with the caspase inhibitor p35, excessive nonautonomous cell proliferation was observed. Significantly, wg signaling is necessary and, at least in some cells, also sufficient for mitogenesis under these conditions. Finally, we provide evidence that the DIAP1 antagonists reaper and hid can activate the JNK pathway and that this pathway is required for inducing wg and cell proliferation. These findings support a model where apoptotic cells activate signaling cascades for compensatory proliferation.     

A comparison of programmed cell death between species

Key components of the programmed cell death pathway are conserved between Caenorhabditis elegans, Drosophila melanogaster and humans. The search for additional homologs has been facilitated by the availability of the entire genomic sequence for each of these organisms.     

The zinc finger homeodomain-2 gene of Drosophila controls Notch targets and regulates apoptosis in the tarsal segments

The development of the Drosophila leg is a good model to study processes of pattern formation, cell death and segmentation. Such processes require the coordinate activity of different genes and signaling pathways that progressively subdivide the leg territory into smaller domains. One of the main pathways needed for leg development is the Notch pathway, required for determining the proximo-distal axis of the leg and for the formation of the joints that separate different leg segments. The mechanisms required to coordinate such events are largely unknown. We describe here that the zinc finger homeodomain-2 (zfh-2) gene is highly expressed in cells that will form the leg joints and needed to establish a correct size and pattern in the distal leg. There is an early requirement of zfh-2 to establish the correct proximo-distal axis, but zfh-2 is also needed at late third instar to form the joint between the fourth and fifth tarsal segments. The expression of zfh-2 requires Notch activity but zfh-2 is necessary, in turn, to activate Notch targets such as Enhancer of split and big brain. zfh-2 is controlled by the Drosophila activator protein 2 gene and regulates the late expression of tarsal-less. In the absence of zfh-2 many cells ectopically express the pro-apoptotic gene head involution defective, activate caspase-3 and are positive for acridine orange, indicating they undergo apoptosis. Our results demonstrate the key role of zfh-2 in the control of cell death and Notch signaling during leg development.     

Detection of apoptosis by terminal deoxynucleotidyl transferase-mediated dUTP nick-end labeling and acridine orange in Drosophila embryos and adult male gonads

In Drosophila, vast numbers of cells undergo apoptosis during normal development. In addition, excessive apoptosis can be induced in response to a variety of stress or injury paradigms, including DNA damage, oxidative stress, nutrient deprivation, unfolded proteins and mechanical tissue damage. Two of the most commonly used methods to label apoptotic cells in Drosophila are terminal deoxynucleotidyl transferase-mediated dUTP nick-end labeling (TUNEL) for fixed tissues and acridine orange (AO) staining for live embryos or tissues. Here, we describe protocols for labeling apoptotic cells in Drosophila embryos and adult male gonads. Slightly modified protocols can also be applied for other Drosophila tissues. The AO protocol is quick, simple and allows real-time imaging of doomed cells in live tissues. However, it is difficult to combine with conventional counterstains or Ab labeling. On the other hand, this functionality is readily afforded by the TUNEL protocol, which permits the detection of apoptotic cells in fixed tissues. These staining procedures can be completed in 1-2 d.     

Cytochrome c-d regulates developmental apoptosis in the Drosophila retina

The role of cytochrome c (Cyt c) in caspase activation has largely been established from mammalian cell-culture studies, but much remains to be learned about its physiological relevance in situ. The role of Cyt c in invertebrates has been subject to considerable controversy. The Drosophila genome contains distinct cyt c genes: cyt c-p and cyt c-d. Loss of cyt c-p function causes embryonic lethality owing to a requirement of the gene for mitochondrial respiration. By contrast, cyt c-d mutants are viable but male sterile. Here, we show that cyt c-d regulates developmental apoptosis in the pupal eye. cyt c-d mutant retinas show a profound delay in the apoptosis of superfluous interommatidial cells and perimeter ommatidial cells. Furthermore, there is no apoptosis in mutant retinal tissues for the Drosophila homologues of apoptotic protease-activating factor 1 (Ark) and caspase 9 (Dronc). In addition, we found that cyt c-d--as with ark and dronc-regulates scutellar bristle number, which is known to depend on caspase activity. Collectively, our results indicate a role of Cyt c in caspase regulation of Drosophila somatic cells.     

Development of a new class of aromatase inhibitors: design, synthesis and inhibitory activity of 3-phenylchroman-4-one (isoflavanone) derivatives

Aromatase (CYP19) catalyzes the aromatization reaction of androgen substrates to estrogens, the last and rate-limiting step in estrogen biosynthesis. Inhibition of aromatase is a new and promising approach to treat hormone-dependent breast cancer. We present here the design and development of isoflavanone derivatives as potential aromatase inhibitors. Structural modifications were performed on the A and B rings of isoflavanones via microwave-assisted, gold-catalyzed annulation reactions of hydroxyaldehydes and alkynes. The in vitro aromatase inhibition of these compounds was determined by fluorescence-based assays utilizing recombinant human aromatase (baculovirus/insect cell-expressed). The compounds 3-(4-phenoxyphenyl)chroman-4-one (1h), 6-methoxy-3-phenylchroman-4-one (2a) and 3-(pyridin-3-yl)chroman-4-one (3b) exhibited potent inhibitory effects against aromatase with IC(50) values of 2.4 μM, 0.26 μM and 5.8 μM, respectively. Docking simulations were employed to investigate crucial enzyme/inhibitor interactions such as hydrophobic interactions, hydrogen bonding and heme iron coordination. This report provides useful information on aromatase inhibition and serves as a starting point for the development of new flavonoid aromatase inhibitors.     

Drosophila IAP antagonists form multimeric complexes to promote cell death

Intraflagellar transport (IFT) machinery mediates the bidirectional movement of cargos that are required for the assembly and maintenance of cilia. However, little is known about how IFT is regulated in vivo. In this study, we show that the small guanosine triphosphatase (GTPase) adenosine diphosphate ribosylation factor–like protein 13 (ARL-13) encoded by the Caenorhabditis elegans homologue of the human Joubert syndrome causal gene ARL13B, localizes exclusively to the doublet segment of the cilium. arl-13 mutants have shortened cilia with various ultrastructural deformities and a disrupted association between IFT subcomplexes A and B. Intriguingly, depletion of ARL-3, another ciliary small GTPase, partially suppresses ciliogenesis defects in arl-13 mutants by indirectly restoring binding between IFT subcomplexes A and B. Rescue of arl-13 mutants by ARL-3 depletion is mediated by an HDAC6 deacetylase-dependent pathway. Thus, we propose that two conserved small GTPases, ARL-13 and ARL-3, coordinate to regulate IFT and that perturbing this balance results in cilia deformation.