title: Expression of genes encoding antimicrobial and bradykinin-related peptides in skin of the stream brown frog Rana sakuraii.
abstract: Peptidomic analysis of an extract of the skin of the stream brown frog Rana sakuraii Matsui and Matsui, 1990 led to the isolation of a C-terminally alpha-amidated peptide (VR-23; VIGSILGALASGLPTLISWIKNR x NH2) with broad-spectrum antimicrobial activity that shows structural similarity to the bee venom peptide, melittin together with two peptides belonging to the temporin family (temporin-1SKa; FLPVILPVIGKLLNGIL x NH2 and temporin-1SKb; FLPVILPVIGKLLSGIL x NH2), and peptides whose primary structures identified them as belonging to the brevinin-2 (2 peptides) and ranatuerin-2 (1 peptide) families. Using a forward primer that was designed from a conserved region of the 5'-untranslated regions of Rana temporaria preprotemporins in a 3'-RACE procedure, a cDNA clone encoding preprotemporin-1SKa was prepared from R. sakuraii skin total RNA. Further preprotemporin cDNAs encoding temporin-1SKc (AVDLAKIANIAN KVLSSL F x NH2) and temporin-1SKd (FLPMLAKLLSGFL x NH2) were obtained by RT-PCR. Unexpectedly, the 3'-RACE procedure using the same primer led to amplification of a cDNA encoding a preprobradykinin whose signal peptide region was identical to that of preprotemporin-1SKa except for the substitution Ser18-->Asn. R. sakuraii bradykinin ([Arg0,Leu1,Thr6,Trp8] BK) was 28-fold less potent than mammalian BK in effecting B2 receptor-mediated relaxation of mouse trachea and the des[Arg0] derivative was only a weak partial agonist. The evolutionary history of the Japanese brown frogs is incompletely understood but a comparison of the primary structures of the R. sakuraii dermal peptides with those of Tago's brown frog Rana tagoi provides evidence for a close phylogenetic relationship between these species.
authors.affiliation: Department of Biology, Faculty of Science, Toho University, 2-2-1 Miyama Funabashi, Chiba 274-8510, Japan.
authors.forename: Hiroe
authors.initials: H
authors.lastname: Suzuki
authors.forename: Shawichi
authors.initials: S
authors.lastname: Iwamuro
authors.forename: Aya
authors.initials: A
authors.lastname: Ohnuma
authors.forename: Laurent
authors.initials: L
authors.lastname: Coquet
authors.forename: Jérôme
authors.initials: J
authors.lastname: Leprince
authors.forename: Thierry
authors.initials: T
authors.lastname: Jouenne
authors.forename: Hubert
authors.initials: H
authors.lastname: Vaudry
authors.forename: Christopher K
authors.initials: CK
authors.lastname: Taylor
authors.forename: Peter W
authors.initials: PW
authors.lastname: Abel
authors.forename: J Michael
authors.initials: JM
authors.lastname: Conlon
mesh_terms: D020121:5' Untranslated Regions; D000595:Amino Acid Sequence; D029845:Amphibian Proteins / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification; D000818:Animals; D023181:Antimicrobial Cationic Peptides / Q000737:chemistry / Q000235:genetics* / Q000302:isolation & purification; D001920:Bradykinin / Q000737:chemistry / Q000235:genetics* / Q000302:isolation & purification / Q000494:pharmacology; D003001:Cloning, Molecular; D018076:DNA, Complementary / Q000235:genetics; D005260:Female; D015870:Gene Expression; D066298:In Vitro Techniques; D008297:Male; D008555:Melitten / Q000737:chemistry / Q000235:genetics; D051379:Mice; D008969:Molecular Sequence Data; D009126:Muscle Relaxation / Q000187:drug effects; D010455:Peptides / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification; D011506:Proteins / Q000737:chemistry / Q000235:genetics; D011898:Ranidae / Q000235:genetics* / Q000378:metabolism; D017386:Sequence Homology, Amino Acid; D012867:Skin / Q000378:metabolism
chemical_list: D020121:5' Untranslated Regions; D029845:Amphibian Proteins; D023181:Antimicrobial Cationic Peptides; D018076:DNA, Complementary; D010455:Peptides; D011506:Proteins; C121230:ranatuerin; C104501:temporin; D008555:Melitten; D001920:Bradykinin