drosulfakinin-ii and drosulfakinin-1

The ability to design agonists that target peptide signaling is a strategy to delineate underlying mechanisms and influence biology. A sequence that uniquely characterizes a peptide provides a distinct site to generate novel agonists. Drosophila melanogaster sulfakinin encodes non-sulfated drosulfakinin I (nsDSK I; FDDYGHMRF-NH2) and nsDSK II (GGDDQFDDYGHMRF-NH2). Drosulfakinin is typical of sulfakinin precursors, which are conserved throughout invertebrates. Non-sulfated DSK II is structurally related to DSK I, however, it contains a unique 5-residue N-terminal extension; drosulfakinins signal through G-protein coupled receptors, DSK-R1 and DSK-R2. Drosulfakinin II distinctly influences adult and larval gut motility and larval locomotion; yet, its structure-activity relationship was unreported. We hypothesized substitution of an N-terminal extension residue may alter nsDSK II activity. By targeting the extension we identified, not unexpectedly, analogs mimicking nsDSK II, yet, surprisingly, we also discovered novel agonists with increased (super) and opposite (protean) effects. We determined [A3] nsDSK II increased larval gut contractility rather than, like nsDSK II, decrease it. [N4] nsDSK II impacted larval locomotion, although nsDSK II was inactive. In adult gut, [A1] nsDSK II, [A2] nsDSKII, and [A3] nsDSK II mimicked nsDSK II, and [A4] nsDSK II and [A5] nsDSK II were more potent; [N3] nsDSK II and [N4] nsDSK II mimicked nsDSK II. This study reports nsDSK II signals through DSK-R2 to influence gut motility and locomotion, identifying a novel role for the N-terminal extension in sulfakinin biology and receptor activation; it also led to the discovery of nsDSK II structural analogs that act as super and protean agonists.

Topics: Amino Acid Sequence; Animals; Drosophila melanogaster; Drosophila Proteins; Intercellular Signaling Peptides and Proteins; Larva; Locomotion; Neuropeptides; Oligopeptides; Peptides; Receptors, Cholecystokinin; Receptors, G-Protein-Coupled; Signal Transduction; Structure-Activity Relationship

Neuropeptides regulate the frequency of heart contractions. Drosophila melanogaster sulfakinin (drosulfakinin) encodes FDDYGHMRFamide, DSK I, and GGDDQFDDYGHMRFamide, DSK II. Invertebrate sulfakinins are structurally and functionally related to vertebrate cholecystokinins. Naturally-occurring drosulfakinins contain a sulfated or nonsulfated tyrosine and are designated sDSK I, sDSK II, nsDSK I, and nsDSK II. We developed a novel neural-cardiovascular preparation and investigated mechanisms regulating the effect of sulfakinins on D. melanogaster heart. We established the preparation in larva, pupa, and adult to examine plasticity in neural regulation of cardiovascular parameters. We discovered sDSK I increased the frequency of larval, pupal, and adult heart contractions; nsDSK I only increased the frequency of larval contractions, not pupal or adult. We also discovered sDSK II and nsDSK II increased the frequency of larval and adult contractions, not pupal. This is the first report of nonsulfated sulfakinin activity on heart, and sulfakinin activity examined in 3 developmental stages within the same animal species. Our data demonstrate a role for plasticity in the effects of sulfakinins on heart contractions, and suggest multiple mechanisms are involved.

Topics: Animals; Drosophila melanogaster; Intercellular Signaling Peptides and Proteins; Myocardial Contraction; Neuropeptides; Oligopeptides; Peptides

Sulfakinins are myoactive peptides and antifeedant factors. Naturally occurring drosulfakinin I (DSK I; FDDYGHMRFNH(2)) and drosulfakinin II (DSK II; GGDDQFDDYGHMRFNH(2)) contain sulfated or nonsulfated tyrosine. We discovered sDSK II and nsDSK II influenced Drosophila melanogaster larval odor preference. However, sDSK I, nsDSK I, MRFNH(2), and saline did not influence odor preference. We discovered sDSK I and nsDSK I influenced larval locomotion. However, sDSK II, nsDSK II, MRFNH(2), and saline did not influence locomotion. Our novel data suggest distinct mechanisms underlie the effects of DSK I and DSK II peptides on odor preference and locomotion, parameters important to many facets of animal survival.

Topics: Amino Acid Sequence; Animals; Drosophila melanogaster; Drosophila Proteins; Intercellular Signaling Peptides and Proteins; Larva; Locomotion; Molecular Sequence Data; Neuropeptides; Odorants; Oligopeptides; Peptides

Sulfakinins, which are satiety factors in invertebrates, have previously been shown to inhibit feeding in the German cockroach and desert locust. This study examines the occurrence of sulfakinin immunoreactivity and the role of sulfakinin as a feeding satiety factor in the black blow fly, Phormia regina. Specifically, this study examines the effect of sulfakinin on two of the blow fly's nutrient requirements (i.e., carbohydrates and proteins). We observed sulfakinin immunoreactive cells in the brains of both male and female flies. We found that drosulfakinin I (DrmSKI, FDDY[SO(3)H]GHMRFa) significantly inhibited carbohydrate feeding by 44% at the most effective dose (10 nmol) in female flies. Statistically, there was no significant effect on males; however, injections of 10 nmol DrmSKI reduced carbohydrate feeding by 34% compared to the sham. Drosulfakinin had no effect on protein feeding and no significant inhibition was detected in females or males. The results of this study lend further support to the idea that carbohydrate and protein feeding are regulated by separate control mechanisms, especially in Calliphoridae.

Topics: Animals; Brain; Dietary Carbohydrates; Dietary Proteins; Diptera; Feeding Behavior; Female; Ganglia; Intercellular Signaling Peptides and Proteins; Neuropeptides; Oligopeptides; Peptides; Protein Transport; Satiety Response; Sex Characteristics

The spatial and temporal distribution of three peptides, DSK I, DSK II, and DSK 0, encoded by the Drosophila melanogaster drosulfakinin (Dsk) gene, have been examined in the central nervous system. DSK I and DSK II have a -RFamide C-terminus and are structurally similar to sulfakinin peptides; in contrast, DSK 0 contains -SFamide and is not structurally similar to sulfakinins. Antisera specificities were determined by the design of the antigens and confirmed by dot blot analysis and preincubation with peptides prior to their use in immunocytochemistry. The distribution of immunoreactivity suggests that all three DSK peptides are processed from the polypeptide precursor and expressed in many of the same cells. Expression was observed at all developmental stages with an increase in the level of staining and the number of immunoreactive cells as development progresses. Cells in the brain lobe, optic lobe, subesophageal ganglion, thoracic ganglia, and the eighth abdominal neuromere contain DSK-immunoreactive materials. Immunoreactive fibers project from some cells and extend into the brain and ventral ganglion with regions of extensive arborization. DSK 0 immunoreactivity provides initial evidence for the presence of a -SFamide peptide in neural tissue. The observed expression of DSK-immunoreactive materials throughout development in numerous cells of the central nervous system suggests that DSK peptides may serve as hormones, modulators, or transmitters involved in several functions.

Topics: Amino Acid Sequence; Animals; Antibody Specificity; Antigens; Central Nervous System; Drosophila melanogaster; Fluorescent Antibody Technique, Indirect; Ganglia, Invertebrate; Intercellular Signaling Peptides and Proteins; Molecular Sequence Data; Nerve Fibers; Neuropeptides; Oligopeptides; Peptides

Homologues to the cholecystokinin (CCK)-gastrin peptide family have been cloned from Drosophila. The CCK-like precursor found in Drosophila has been designated drosulfakinin (DSK). Genomic and cDNA clones corresponding to the Drosophila neuropeptide precursor encode for three putative peptides. The three peptides (DSK-0, Asn-Gln-Lys-Thr-Met-Ser-Phe-Gly; DSK-I, Phe-Asp-Asp-Tyr-Gly-His-Met-Arg-Phe-Gly; DSK-II, Gly-Gly-Asp-Asp-Gln-Phe-Asp-Asp-Tyr-Gly-His-Met-Arg-Phe-Gly) are flanked by prohormone processing sites and contain C-terminal glycyl residues, a potential amidation site. Two of the peptides, DSK-I and DSK-II, are homologous to CCK-gastrin peptides. Each of the two homologues include a CCK-gastrin-like C-terminal pentapeptide and a conserved sequence corresponding to the sulfated tyrosine in bioactive CCK. The third peptide encoded by the drosulfakinin precursor represents a novel peptide. In situ tissue hybridization indicates the presence of the transcript in the adult head. Chromosomal localization maps the gene to the third chromosome near 81F.

Topics: Amino Acid Sequence; Animals; Base Sequence; Cholecystokinin; Deoxyribonuclease EcoRI; DNA; DNA Restriction Enzymes; DNA, Recombinant; Drosophila melanogaster; Gastrins; Intercellular Signaling Peptides and Proteins; Molecular Sequence Data; Neuropeptides; Nucleic Acid Hybridization; Oligopeptides; Peptides; Protein Precursors; RNA, Messenger; Sequence Homology, Nucleic Acid