Michael Lehmann

Pipsqueak and GAGA factor act in concert as partners at homeotic and many other loci

The Drosophila GAGA factor (GAF) controls transcription and other chromosome functions by altering chromatin structure. We found that a second GAGA-binding protein of Drosophila, Pipsqueak (Psq), can directly bind to GAF and is associated with GAF in vivo. Genetic interaction studies provide evidence that Psq and GAF act together in the transcriptional activation and silencing of homeotic genes. A complete colocalization of Psq and GAF on polytene interphase chromosomes and mitotic chromosomes suggests that the two proteins cooperate as general partners not only at homeotic loci, but also at hundreds of other chromosomal sites.

Lipin Is a Central Regulator of Adipose Tissue Development and Function in Drosophila melanogaster

ABSTRACT Lipins are evolutionarily conserved proteins found from yeasts to humans. Mammalian and yeast lipin proteins have been shown to control gene expression and to enzymatically convert phosphatidate to diacylglycerol, an essential precursor in triacylglcerol (TAG) and phospholipid synthesis. Loss of lipin 1 in the mouse, but not in humans, leads to lipodystrophy and fatty liver disease. Here we show that the single lipin orthologue of Drosophila melanogaster (dLipin) is essential for normal adipose tissue (fat body) development and TAG storage. dLipin mutants are characterized by reductions in larval fat body mass, whole-animal TAG content, and lipid droplet size. Individual cells of the underdeveloped fat body are characterized by increased size and ultrastructural defects affecting cell nuclei, mitochondria, and autophagosomes. Under starvation conditions, dLipin is transcriptionally upregulated and functions to promote survival. Together, these data show that dLipin is a central player in lipid and energy metabolism, and they establish Drosophila as a genetic model for further studies of conserved functions of the lipin family of metabolic regulators.

HIV-1 Nef upregulates CCL2/MCP-1 expression in astrocytes in a myristoylation- and calmodulin-dependent manner

HIV-associated dementia (HAD) correlates with infiltration of monocytes into the brain. The accessory HIV-1 negative factor (Nef) protein, which modulates several signaling pathways, is constitutively present in persistently infected astroctyes. We demonstrated that monocytes responded with chemotaxis when subjected to cell culture supernatants of nef-expressing astrocytic U251MG cells. Using a protein array, we identified CC chemokine ligand 2/monocyte chemotactic protein-1 (CCL2/MCP-1) as a potential chemotactic factor mediating this phenomenon. CCL2/MCP-1 upregulation by Nef was further confirmed by ribonuclease protection assay, RT-PCR and ELISA. By applying neutralizing antibodies against CCL2/MCP-1 and using CCR2-deficient monocytes, we confirmed CCL2/MCP-1 as the exclusive factor secreted by nef-expressing astrocytes capable of attracting monocytes. Additionally, we showed that Nef-induced CCL2/MCP-1 expression depends on the myristoylation moiety of Nef and requires functional calmodulin. In summary, we suggest that Nef-induced CCL2/MCP-1 expression in astrocytes contributes to infiltration of monocytes into the brain, and thereby to progression of HAD.

Fork head controls the timing and tissue selectivity of steroid-induced developmental cell death

Cell death during Drosophila melanogaster metamorphosis is controlled by the steroid hormone 20-hydroxyecdysone (20E). Elements of the signaling pathway that triggers death are known, but it is not known why some tissues, and not others, die in response to a particular hormone pulse. We found that loss of the tissue-specific transcription factor Fork head (Fkh) is both required and sufficient to specify a death response to 20E in the larval salivary glands. Loss of fkh itself is a steroid-controlled event that is mediated by the 20E-induced BR-C gene, and that renders the key death regulators hid and reaper hormone responsive. These results implicate the D. melanogaster FOXA orthologue Fkh with a novel function as a competence factor for steroid-controlled cell death. They explain how a specific tissue is singled out for death, and why this tissue survives earlier hormone pulses. More generally, they suggest that cell identity factors like Fkh play a pivotal role in the normal control of developmental cell death.

Ecdysteroid receptors of the blowfly Calliphora vicina: Partial purification and characterization of ecdysteroid binding

A macromolecule with high affinity for the ecdysteroid analogue ponasterone A was isolated from nuclei of larvae of the blowfly Calliphora vicina. The ecdysteroid-binding molecule revealed characteristics of the moulting hormone receptor. It was sensitive towards protease but not towards nucleases. The nuclear protein had a limited binding capacity (0.2 pmol ponasterone A/mg protein), showed hormone analogue specificity and high affinity for ecdysteroids. Enzyme activities were present in the nuclear extract that metabolized ecdysteroids and thereby interfered with the binding assay. After their removal by DEAE-cellulose chromatography the ecdysteroid receptor preparation was stable at 20 degrees C for hours. This allowed a reliable determination of dissociation constants at equilibrium conditions. The hormone receptor complex had a KD of 1 nM, 30 nM, and 2000 nM with ponasterone A, 20-hydroxyecdysone, and ecdysone, respectively. The apparent molecular mass of the ecdysteroid receptor was 105,000 as determined by gel filtration.

FOXO-independent suppression of programmed cell death by the PI3K/Akt signaling pathway in Drosophila

Signaling through the PI3K/Akt/FOXO pathway plays an important role in vertebrates in protecting cells from programmed cell death. PI3K and Akt have been similarly shown to be involved in survival signaling in the invertebrate model organism Drosophila. However, it is not known whether PI3K and Akt execute this function by controlling a pro-apoptotic activity of Drosophila FOXO. In this study, we show that elevated signaling through PI3K and Akt can prevent developmentally controlled death in the salivary glands of the fruit fly. We further show that Drosophila FOXO is not required for normal salivary gland death and that the rescue of salivary gland death by PI3K occurs independent of FOXO. These results give support to the notion that FOXOs have acquired pro-apoptotic functions after separation of the vertebrate and invertebrate lineages.

Drosophila Lipin interacts with insulin and TOR signaling pathways in the control of growth and lipid metabolism

ABSTRACT Lipin proteins have key functions in lipid metabolism, acting as both phosphatidate phosphatases (PAPs) and nuclear regulators of gene expression. We show that the insulin and TORC1 pathways independently control functions of Drosophila Lipin (dLipin). Reduced signaling through the insulin receptor strongly enhanced defects caused by dLipin deficiency in fat body development, whereas reduced signaling through TORC1 led to translocation of dLipin into the nucleus. Reduced expression of dLipin resulted in decreased signaling through the insulin-receptor-controlled PI3K–Akt pathway and increased hemolymph sugar levels. Consistent with this, downregulation of dLipin in fat body cell clones caused a strong growth defect. The PAP but not the nuclear activity of dLipin was required for normal insulin pathway activity. Reduction of other enzymes of the glycerol-3 phosphate pathway affected insulin pathway activity in a similar manner, suggesting an effect that is mediated by one or more metabolites associated with the pathway. Taken together, our data show that dLipin is subject to intricate control by the insulin and TORC1 pathways, and that the cellular status of dLipin impacts how fat body cells respond to signals relayed through the PI3K–Akt pathway. Summary: The insulin receptor and TORC1 pathways independently control cytoplasmic and nuclear functions of Drosophila Lipin, and Lipin, in turn, modulates cellular responses to growth factor signaling.

Endocrine and physiological regulation of neutral fat storage in Drosophila

After having revolutionized our understanding of the mechanisms of animal development, Drosophila melanogaster has more recently emerged as an equally valid genetic model in the field of animal metabolism. An increasing number of studies have revealed that many signaling pathways that control metabolism in mammals, including pathways controlled by nutrients (insulin, TOR), steroid hormone, glucagon, and hedgehog, are functionally conserved between mammals and Drosophila. In fact, genetic screens and analyses in Drosophila have identified new players and filled in gaps in the signaling networks that control metabolism. This review focuses on data that show how these networks control the formation and breakdown of triacylglycerol energy stores in the fat tissue of Drosophila.

Roles of the FOXA transcription factor Fork head in autophagic developmental cell death.

Fork head (Fkh) is the only fruit fly ortholog of the FOXA subgroup of FOX transcription factors. Recent studies on the role of Fkh in autophagic cell death during Drosophila metamorphosis identified Fkh as a competence factor for steroid-induced cell death. Microarray analyses suggest that, in addition to controlling essential apoptosis genes, Fkh controls autophagic and cell survival pathways. The data support a model in which Fkh promotes cell growth and survival by coordinating the cell’s ability to respond to death and autophagy-controlling signals. Addendum to: Liu Y, Lehmann M. Genes and biological processes controlled by the Drosophila FOXA orthologue Fork head. Insect Mol Biol 2008; 17:91-101.