title: Molecular cloning, expression and identification of the promoter regulatory region for the neuropeptide trissin in the nervous system of the silkmoth Bombyx mori.
abstract: Trissin has recently been identified as a conserved insect neuropeptide, but its cellular expression and function is unknown. We detected the presence of this neuropeptide in the silkworm Bombyx mori using in silico search and molecular cloning. In situ hybridisation was used to examine trissin expression in the entire central nervous system (CNS) and gut of larvae, pupae and adults. Surprisingly, its expression is restricted to only two pairs of small protocerebral interneurons and four to five large neurons in the frontal ganglion (FG). These neurons were further characterised by subsequent multiple staining with selected antibodies against insect neuropeptides. The brain interneurons innervate edges of the mushroom bodies and co-express trissin with myoinhibitory peptides (MIP) and CRF-like diuretic hormones (CRF-DH). In the FG, one pair of neurons co-express trissin with calcitonin-like diuretic hormone (CT-DH), short neuropeptide F (sNPF) and MIP. These neurons innervate the brain tritocerebrum and musculature of the anterior midgut. The other pair of trissin neurons in the FG co-express sNPF and project axons to the tritocerebrum and midgut. We also used the baculovirus expression system to identify the promoter regulatory region of the trissin gene for targeted expression of various molecular markers in these neurons. Dominant expression of trissin in the FG indicates its possible role in the regulation of foregut-midgut contractions and food intake.
authors.affiliation: Institute of Zoology, Slovak Academy of Sciences, Dúbravská cesta 9, 845 06, Bratislava, Slovakia.
authors.forename: Ladislav
authors.initials: L
authors.lastname: Roller
authors.affiliation: Institute of Zoology, Slovak Academy of Sciences, Dúbravská cesta 9, 845 06, Bratislava, Slovakia.
authors.forename: Daniel
authors.initials: D
authors.lastname: Čižmár
authors.affiliation: Institute of Zoology, Slovak Academy of Sciences, Dúbravská cesta 9, 845 06, Bratislava, Slovakia.
authors.forename: Zuzana
authors.initials: Z
authors.lastname: Gáliková
authors.affiliation: Institute of Zoology, Slovak Academy of Sciences, Dúbravská cesta 9, 845 06, Bratislava, Slovakia.
authors.forename: Branislav
authors.initials: B
authors.lastname: Bednár
authors.affiliation: Institute of Zoology, Slovak Academy of Sciences, Dúbravská cesta 9, 845 06, Bratislava, Slovakia.
authors.forename: Ivana
authors.initials: I
authors.lastname: Daubnerová
authors.affiliation: Institute of Zoology, Slovak Academy of Sciences, Dúbravská cesta 9, 845 06, Bratislava, Slovakia. dusan.zitnan@savba.sk.
authors.forename: Dušan
authors.initials: D
authors.lastname: Žitňan
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D012831:Bombyx / Q000235:genetics*; D002490:Central Nervous System / Q000378:metabolism*; D003001:Cloning, Molecular; D018076:DNA, Complementary / Q000235:genetics; D017952:Ganglia, Invertebrate / Q000378:metabolism; D005786:Gene Expression Regulation; D049452:Green Fluorescent Proteins / Q000378:metabolism; D007150:Immunohistochemistry; D017403:In Situ Hybridization; D007301:Insect Hormones / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D007814:Larva / Q000378:metabolism; D009474:Neurons / Q000166:cytology / Q000378:metabolism; D009479:Neuropeptides / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D011401:Promoter Regions, Genetic / Q000235:genetics*
chemical_list: D018076:DNA, Complementary; D007301:Insect Hormones; D009479:Neuropeptides; C485184:enhanced green fluorescent protein; D049452:Green Fluorescent Proteins
keywords: Gut; Immunohistochemistry; In situ hybridisation; Neuron; Neuropeptide