Alternative splicing directs the expression of two D2 dopamine receptor isoforms

Dopamine receptors are classified into D1 and D2 subtypes on the basis of their pharmacological properties and the intracellular responses they mediate. The cerebral D2 dopamine receptor is the target of drugs used to alleviate the main symptoms of schizophrenia. Although it is considered to be a single molecular entity, there is evidence that multiple D2-receptor subtypes exist. A complementary DNA encoding a D2 receptor has recently been cloned and the deduced 415-amino-acid sequence indicates that it belongs to the large superfamily of receptors coupled to G proteins, and that its topology consists of seven transmembrane domains. In this family, the genes are frequently without introns and each is believed to encode a unique polypeptide product. Here we show that the gene for the D2 receptor produces two receptor isoforms by alternative messenger RNA splicing, providing a route to receptor diversity in this family. One isoform corresponds to the D2(415) receptor, but the second contains an additional sequence encoding a 29-amino-acid fragment, defining a novel D2(444) receptor isoform. Expression of the two isoforms is tissue-specific, and both are regulated by guanyl nucleotides. As the extra sequence is located within a putative cytoplasmic loop that binds to G proteins, the two isoforms might interact with different G proteins and thereby initiate distinct intracellular signals.     

Mitochondrial DNA variation in social wasps (Hymenoptera,Vespidae)

Summary Patterns of restriction fragment length polymorphisms (RFLP) of European Vespinae were more similar within genera than between them. Distance trees were constructed that support the hypothesis of monophyly of the generaVespula andDolichovespula. Within the genusVespula, V. germanica was more closely related toV. rufa than toV. vulgaris. The position of the genusVespa remained uncertain due to the precision limits of the RFLP technique.     

Cryopreservation of Drosophila melanogaster embryos

There is an urgent need to preserve the ever-increasing number (greater than 30,000) of different genetic strains of D. melanogaster that are maintained in national and international stock centres and in the laboratories of individual investigators. In all cases, the stocks are maintained as adult populations and require transfer to fresh medium every two to four weeks. This is not only costly in terms of materials, labour and space, but unique strains are vulnerable to accidental loss, contamination, and changes in genotype that can occur during continuous culture through mutation, genetic drift or selection. Although cryopreservation of Drosophila germ-plasm would be an enormous advantage, many attempts using conventional procedures have been unsuccessful. D. melanogaster embryos are refractory to conventional cryopreservation procedures because of the contravening conditions required to minimize mortality resulting from both intracellular ice formation and chilling injury at subzero temperatures. To overcome these obstacles, we have developed a vitrification procedure that precludes intracellular ice formation so that the embryos can be cooled and warmed at ultra-rapid rates to minimize chilling injury, and have recovered viable embryos following storage in liquid nitrogen. In a series of 53 experiments, a total of 3,711 larvae emerged from 17,280 eggs that were cooled in liquid nitrogen (18.4 +/- 8.8%). Further, using a subset from this population, approximately 3% of the surviving larvae (24/800) developed into adults. These adults were fertile and produced an F1 generation.     

Transmembrane movements of lipids

Summary Membranes allow the rapid passage of uncharged lipids. Phospholipids on the other hand diffuse very slowly from one monolayer to another with a half-time of several hours. This slow spontaneous movement in a pure lipid bilayer can be selectively modulated in biological membranes by intrinsic proteins. In microsomes, and probably in bacterial membranes, non-specific phospholipid flippases allow the rapid redistribution of newly synthesized phospholipids. In eukaryotic plasma membranes, aminophospholipid translocase selectively pumps phosphatidylserine (PS) and phosphatidylethanolamine (PE) from the outer to the inner leaflet and establishes a permanent lipid asymmetry. The discovery of an aminophospholipid translocase in chromaffin granules proves that eukaryotic organelles may also contain lipid translocators.     

Competition for antigen presentation in living cells involves exchange of peptides bound by class II MHC molecules

T cells recognize foreign proteins as peptides bound to self molecules encoded by the major histocompatibility complex (MHC). The kinetics of interaction between purified class II MHC molecules and peptides is unusual, in that the rate of association is very slow, but once formed, the complexes are extremely stable. This raises the question of how the antigen-presenting cell provides a sufficient number of free MHC binding sites to ensure T cell immunity. We present results suggesting that an exchange of peptide in MHC binding sites may take place under physiological conditions.     

Postmolt cuticle growth in a cockroach: in vitro deposition of multilamellate and circadian-like layered endocuticle

Summary Pieces of the hind legs of freshly moltedBlaberus craniifer adults were cultivated in vitro. The deposition of multilamellate and circadian-like layered endocuticle was observed. In multilamellate cuticle, the number of double lamellae was frequently much greater than the number of days in culture. In circadian-like layered cuticle, the number of double layers corresponded to the numbers of days in culture, or it was smaller.I am indebted to Mrs Hassenrück, for excellent assistance. — Supported by the Deutsche Forschungsgemeinschaft (We 389/12-2).     

Esterases of the flounder (Platichthys flesus,Pleuronectidae, Teleostei): Development of an identification protocol using starch gel electrophoresis and characterization of loci

Summary Fish esterases are among the most difficult enzymes to identify using starch gel electrophoresis because of the many loci that are simultaneously active, and especially because of duplication phenomena, satellite bands, and stain trails. In an attempt to simplify and clarify electropherograms, various staining and inhibitory methods were tested on esterases from the flounderPlatichthys flesus. A range of migration and staining buffers and substrates were used, as well as chemical inhibition and heat inactivation. A combination of the methods made it possible to distinguish and characterize the five presumed esterase loci.