Marie-Marthe Suner

Functional expression of a locust tyramine receptor in murine erythroleukaemia cells

The LCR/MEL system (Locus Control Region/Murine Erythroleukaemia cells) was employed to express and characterize the Locusta migratoria tyramine receptor (TyrLoc), an insect G protein‐coupled receptor. Functional agonist‐dependent responses were recorded in stable, tyramine receptor expressing cell clones (MEL‐TyrLoc). Tyramine elicited a dose‐dependent increase of cytosolic Ca2+‐ions and an attenuation of forskolin‐induced cyclic adenosine monophosphate (AMP) production. Octopamine was shown to be a weak agonist for both responses. In addition, yohimbine proved to be a potent tyramine receptor antagonist. This study reports the first application of the LCR/MEL expression system in functional assays for G protein‐coupled receptors and therefore expands the capabilities of this system by exploiting the functionality of the signal transduction pathways.

Identification of a polymorphic ryanodine receptor gene from Heliothis virescens (Lepidoptera: noctuidae).

cDNAs encoding the C-terminal 1172 amino acids of a ryanodine receptor (RyR) from the lepidopteran pest Heliothis virescens (Hv-RyR) have been cloned and characterised. Sequence comparisons, organisational studies on corresponding genomic regions and a genetic segregation analysis provide evidence for two polymorphic alleles of the Hv-RyR locus. Comparison of the Hv-RyR C-terminal amino acid sequence with equivalent regions of other RyRs reveals a high level of overall amino acid homology (74% identity with D. melanogaster and between 47.9 and 50.1% with vertebrate isoforms). Homologies are however not uniformly distributed, though regions of high and low similarity are consistent with patterns in other RyR isoforms. The structural similarity of Hv-RyR with other RyRs is also indicated by comparison of hydropathy profiles and other previously described functional domains. Such results are consistent with this region of Hv-RyR containing the Ca(2+) channel itself and being intimately involved in RyR regulation. Potential uses of the cDNAs described in the discovery and development of novel ryanodine like insecticides are discussed.

Glucocorticoid-inducible gene expression vectors for use in Drosophila melanogaster Schneider 2 cells

Inducible, vector‐based, expression systems that allow fine control of transgene expression are gaining more and more use in fundamental research as well as in therapeutic applications. In an effort to develop a tightly regulated heterologous expression system for Drosophila Schneider 2 cells, three different inducible reporter constructs were compared. These comprised six copies of the glucocorticoid response element fused to one of three distinct types of Drosophila gene promoters: (1) a TATA‐box containing, (2) a TATA‐less and (3) a bidirectional core sequence. These were fused to a luciferase reporter gene. The promoter constructs displayed different basal as well as agonist‐induced activities. The implications of the observations made are discussed in the context of promoter properties and of induction of genes that may be studied in Drosophila.