Identification of a fibroblast growth factor-binding protein in Drosophila melanogaster.

As assessed by competitive binding and protein-crosslinking experiments, Drosophila melanogaster cells possess basic fibroblast growth factor (bFGF)-specific binding proteins that are similar to FGF receptors on vertebrate cells in molecular weight and binding affinity; these D. melanogaster cells, however, have no detectable binding proteins for acidic fibroblast growth factor (aFGF). Consistent with the presence of bFGF-specific binding proteins, D. melanogaster cells degrade bFGF but not aFGF. These results indicate the conservation of heparin-binding growth factors and receptors between vertebrates and D. melanogaster.     

The Drosophila melanogaster Cajal body

Cajal bodies (CBs) are nuclear organelles that are usually identified by the marker protein p80-coilin. Because no orthologue of coilin is known in Drosophila melanogaster, we identified D. melanogaster CBs using probes for other components that are relatively diagnostic for CBs in vertebrate cells. U85 small CB-specific RNA, U2 small nuclear RNA, the survival of motor neurons protein, and fibrillarin occur together in a nuclear body that is closely associated with the nucleolus. Based on its similarity to CBs in other organisms, we refer to this structure as the D. melanogaster CB. Surprisingly, the D. melanogaster U7 small nuclear RNP resides in a separate nuclear body, which we call the histone locus body (HLB). The HLB is invariably colocalized with the histone gene locus. Thus, canonical CB components are distributed into at least two nuclear bodies in D. melanogaster. The identification of these nuclear bodies now permits a broad range of questions to be asked about CB structure and function in a genetically tractable organism.     

Nuclear receptors are markers of animal genome evolution

Nuclear hormone receptors form one evolutionary related super-family of proteins, which mediate the interaction between hormones (or other ligands) and gene expression in animals. Early phylogenetic analyses showed two main periods of gene duplication which gave rise to present-day diversity in most animals: one at the origin of the family, and another specifically in vertebrates. Moreover this second period is composed itself by, probably, two rounds of duplication, as proposed by Susumu Ohno at the origin of vertebrates. There are indeed often two, three or four vertebrate orthologs of each invertebrate nuclear receptor, in accordance with this theory. The complete genome of Drosophila melanogaster contains 21 nuclear receptors, compared to 49 in the human genome. In addition, many nuclear receptors have more paralogs in the zebrafish than in mammals, and a genome duplication has been proposed at the origin of ray-finned fishes. Nuclear receptors are a very good model to investigate the dating and functional role of these duplications, since they are dispersed in the genome, allow robust phylogenetic reconstruction, and are functionnaly well characterized, with different adaptations for different paralogs. We illustrate this with examples from differents nuclear receptors and different groups of species.     

Drosophila and vertebrate myb proteins share two conserved regions, one of which functions as a DNA-binding domain

We report the nucleotide sequence of a cDNA clone of the Drosophila melanogaster homologue of c-myb, a member of the class of vertebrate transforming genes encoding nuclear proteins. We predict the mol. wt of the Drosophila myb (D-myb) protein to be 74,000. The D-myb protein contains two clusters of sequences homologous to vertebrate myb proteins, surrounded by sequences lacking homology. These results extend previous evidence for the existence of a D. melanogaster homologue of c-myb and identify two highly conserved and therefore presumably functionally important domains of c-myb proteins. DNA-binding experiments indicate that the NH2-proximal of the two homology regions functions as a DNA-binding domain. Based on the absence of the COOH-proximal homology region in truncated oncogenic derivatives of c-myb it is likely that this homology region encodes a function whose loss is involved in activating the oncogenic potential of c-myb.     

The invertebrate ancestry of endocannabinoid signalling: an orthologue of vertebrate cannabinoid receptors in the urochordate Ciona intestinalis

The G-protein coupled cannabinoid receptors CB(1) and CB(2) are activated by Delta(9)-tetrahydrocannabinol, the psychoactive ingredient of cannabis, and mediate physiological effects of endogenous cannabinoids ('endocannabinoids'). CB(1) genes have been identified in mammals, birds, amphibians and fish, whilst CB(2) genes have been identified in mammals and in the puffer fish Fugu rubripes. Therefore, both CB(1) and CB(2) receptors probably occur throughout the vertebrates. However, cannabinoid receptor genes have yet to be identified in any invertebrate species and the evolutionary origin of cannabinoid receptors is unknown. Here we report the identification of CiCBR, a G-protein coupled receptor in a deuterostomian invertebrate - the urochordate Ciona intestinalis - that is orthologous to vertebrate cannabinoid receptors. The CiCBR cDNA encodes a protein with a predicted length (423 amino-acids) that is the intermediate of human CB(1) (472 amino-acids) and human CB(2) (360-amino-acid) receptors. Interestingly, the protein-coding region of the CiCBR gene is interrupted by seven introns, unlike in vertebrate cannabinoid receptor genes where the protein-coding region is typically intronless. Phylogenetic analysis revealed that CiCBR forms a clade with vertebrate cannabinoid receptors but is positioned outside the CB(1) and CB(2) clades of a phylogenetic tree, indicating that the common ancestor of CiCBR and vertebrate cannabinoid receptors predates a gene (genome) duplication event that gave rise to CB(1)- and CB(2)-type receptors in vertebrates. Importantly, the discovery of CiCBR and the absence of orthologues of CiCBR in protostomian invertebrates such as Drosophila melanogaster and Caenorhabditis elegans indicate that the ancestor of vertebrate CB(1) and CB(2) cannabinoid receptors originated in a deuterostomian invertebrate.     

Powered by gas--a ligand for a fruit fly nuclear receptor

The difficulty in identifying ligands for nuclear hormone receptors remains an obstacle to understanding their function. For example, in the fruit fly Drosophila melanogaster, only one of its nuclear receptors has a known ligand. In this issue of Cell, report that the fruit fly E75 nuclear receptor contains heme in its ligand binding pocket and that the oxidation state of this molecule controls E75 activity. They also show that E75-heme responds to the small diatomic gases, nitric oxide and carbon monoxide. This study sheds light on how heme, gas signaling, and nuclear receptors interact to control metabolic and developmental pathways.     

Identification of the first invertebrate interleukin JAK/STAT receptor, the Drosophila gene domeless.

The JAK/STAT signaling pathway plays important roles in vertebrate development and the regulation of complex cellular processes. Components of the pathway are conserved in Dictyostelium, Caenorhabditis, and Drosophila, yet the complete sequencing and annotation of the D. melanogaster and C. elegans genomes has failed to identify a receptor, raising the possibility that an alternative type of receptor exists for the invertebrate JAK/STAT pathway. Here we show that domeless (dome) codes for a transmembrane protein required for all JAK/STAT functions in the Drosophila embryo. This includes its known requirement for embryonic segmentation and a newly discovered function in trachea specification. The DOME protein has a similar extracellular structure to the vertebrate cytokine class I receptors, although its sequence has greatly diverged. Like many interleukin receptors, DOME has a cytokine binding homology module (CBM) and three extracellular fibronectin-type-III domains (FnIII). Despite its low degree of overall similarity, key amino acids required for signaling in the vertebrate cytokine class I receptors [3] are conserved in the CBM region. DOME is a signal-transducing receptor with most similarities to the IL-6 receptor family, but it also has characteristics found in the IL-3 receptor family. This suggests that the vertebrate families evolved from a single ancestral receptor that also gave rise to dome.     

FGF signaling in flies and worms: more and more relevant to vertebrate biology

FGF signaling in the invertebrate model systems Drosophila melanogaster and Caenorhabditis elegans was initially most obviously involved in cell motility events. More recently, however, FGFs and FGF signaling in these systems have been shown to affect many additional cellular processes. This recent work has shown that the pleiotropies of these FGF receptors resemble those of their vertebrate counterparts, and, in many cases, serve as excellent models for understanding the fundamental molecular mechanisms controlling these events.     

Purification of the vertebrate nuclear pore complex by biochemical criteria

The nuclear pore is a large and complex biological machine, mediating all signal-directed transport between the nucleus and the cytoplasm. The vertebrate pore has a mass of ∼120 million daltons or 30 times the size of a ribosome. The large size of the pore, coupled to its tight integration in the nuclear lamina, has hampered the isolation of pore complexes from vertebrate sources. We have now developed a strategy for the purification of nuclear pores from in vitro assembled annulate lamellae (AL), a cytoplasmic mimic of the nuclear envelope that lacks a lamina, nuclear matrix, and chromatin-associated proteins. We find that purified pore complexes from annulate lamellae contain every nuclear pore protein tested. In addition, immunoblotting reveals the presence of soluble transport receptors and factors known to play important roles in the transport of macromolecules through the pore. While transport factors such as Ran and NTF2 show only transient interaction with the pores, a number of soluble transport receptors, including importin β, show a tight association with the purified pores. In summary, we report that we have purified the vertebrate pore by biochemical criteria; silver staining reveals ∼40–50 distinct protein bands.     

Ionotropic glutamate receptors in Caenorhabditis elegans

Ionotropic glutamate receptors (iGluRs) are an important class of heteromeric ligand-gated receptor complexes that mediate a large portion of the excitatory neurotransmission in the vertebrate CNS. Since the cloning of the first iGluR subunit in 1989, the study of this receptor family has rapidly developed in mammals and expanded to include the study of conserved glutamate receptors in simpler invertebrate systems, including the fruit fly Drosophila melanogaster and the soil nematode Caenorhabditis elegans. These model organisms have enabled the genetic analysis of glutamate receptors in the context of a simpler nervous system and provided new insights into receptor function and regulation. In this review we will focus on recent studies that have used genetic, behavioral, and electrophysiological techniques to study the function of iGluRs in C. elegans.     

The importance of exportin and Ran for nucleocytoplasmic shuttling of the ecdysteroid receptor

Nuclear localization of the ecdysteroid receptor (EcR) is increased in HeLa cells if exportin-1 (CRM1), a predominant carrier for export of proteins and RNA from the nucleus into the cytoplasm, is knocked down by siRNA against exportin. However, knockdown of the small G protein Ran, which is essential for nuclear transport, leads to an arrest of EcR in the cytoplasm, but does not prevent efficient nuclear import of the most important heterodimerization partner of EcR, ultraspiracle (Usp). Like in vertebrate cells, EcR is also distributed heterogeneously in Drosophila melanogaster S2 cells but shifted exclusively to the nucleus, if Usp is present.     

Transrepression of AP-1 by nuclear receptors in Drosophila

Mammalian cell culture studies have shown that several members of the nuclear receptor super family such as glucocorticoid receptor, retinoic acid receptor and thyroid hormone receptor can repress the activity of AP-1 proteins by a mechanism that does not require the nuclear receptor to bind to DNA directly, but that is otherwise poorly understood. Several aspects of nuclear receptor function are believed to rely on this inhibitory mechanism, which is referred to as transrepression. This study presents evidence that nuclear receptor-mediated transrepression of AP-1 occurs in Drosophila melanogaster. In two different developmental situations, embryonic dorsal closure and wing development, several nuclear receptors, including Seven up, Tailless, and Eagle antagonize AP-1. The inhibitory interactions with nuclear receptors are integrated with other modes of AP-1 regulation, such as mitogen-activated protein kinase signaling. A potential role of nuclear receptors in setting a threshold of AP-1 activity required for the manifestation of a cellular response is discussed.     

Retinoids and their targets in vertebrate development

The elaboration of the effect of retinoic acid on limb morphogenesis has prompted renewed investigation into the teratology of retinoic acid treatment, with the hope that such analysis might give insight into mechanisms of vertebrate patterning. Retinoids, their nuclear receptors and their cytoplasmic binding proteins are now known to be deployed throughout development, but the extent to which they are natural agents of morphogenesis remains obscure. The study of retinoic acid receptors may offer molecular insight into gene regulation underlying vertebrate pattern formation.