RESUMO
Dogfish (Squalus acanthias) growth hormone (GH) was identified by cDNA cloning and protein purification from the pituitary gland. Dogfish GH cDNA encoded a prehormone of 210 amino acids (aa). Sequence analysis of purified GH revealed that the prehormone is composed of a signal peptide of 27 aa and a mature protein of 183 aa. Dogfish GH showed 94% sequence identity with blue shark GH, and also showed 37-66%, 26%, and 48-67% sequence identity with GH from osteichtyes, an agnathan, and tetrapods. The site of production was identified through immunocytochemistry to be cells of the proximal pars distalis of the pituitary gland. Dogfish GH stimulates both insulin-like growth factor-I and II mRNA levels in dogfish liver in vitro. The dogfish GH gene consisted of five exons and four introns, the same as in lamprey, teleosts such as cypriniforms and siluriforms, and tetrapods. The 5'-flanking region within 1082 bp of the transcription start site contained consensus sequences for the TATA box, Pit-1/GHF-1, CRE, TRE, and ERE. These results show that the endocrine mechanism for growth stimulation by the GH-IGF axis was established at an early stage of vertebrate evolution, and that the 5-exon-type gene organization might reflect the structure of the ancestral gene for the GH gene family.
Assuntos
Componentes do Gene/genética , Hormônio do Crescimento/genética , Filogenia , Squalus acanthias/genética , Sequência de Aminoácidos , Animais , Sequência de Bases , Clonagem Molecular , Análise por Conglomerados , Primers do DNA/genética , Imuno-Histoquímica , Fígado/metabolismo , Dados de Sequência Molecular , Hipófise/metabolismo , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Análise de Sequência de DNA , Especificidade da EspécieRESUMO
The absolute configurations of the two acid moieties, 12-methyltetradecanoate and 14-methylhexadecanoate, of glycoglycerolipid S365A isolated from Corynebacterium aquaticum were determined by an HPLC analysis after their conversion with the chiral fluorescent labeling reagents, (1S,2S)- and (1R,2R)-2-(2,3-anthracenedicarboximido)cyclohexanol. Both anteiso acids had the S configuration.
Assuntos
Corynebacterium/química , Decanoatos/química , Glicolipídeos/química , Cromatografia Líquida de Alta Pressão/métodos , Glicolipídeos/isolamento & purificação , Indicadores e Reagentes , Conformação MolecularRESUMO
Proopiomelanocortin (POMC) is a precursor for several pituitary hormones including adrenocorticotropin (ACTH), melanocyte-stimulating hormone (MSH) and endorphin (END). Fish POMCs in four taxonomic classes, Cephalaspidomorphi (lampreys), Chondrichthyes (cartilaginous fish), Sarcopterygii (lobe-finned fish), and Actinopterygii (ray-finned fish) have been identified. However, two essential species, ratfish in Chondrichthyes and hagfish in Agnatha, are still missing in the evolutionary image of this molecule. The present study reports analysis of POMC cDNA in the ratfish, Chimaera phantasma, which belongs to another subclass in the Chondrichthyes. Partial cDNA clones were amplified by PCR from single-strand cDNA prepared on total RNA from a complex of pituitary and hypothalamus, and subsequently overlapped to obtain a full-length sequence. Ratfish POMC cDNA consists of 1294bp excluding the poly(A) tail. It encodes a signal peptide of 25 amino acids and POMC of 300 amino acids. gamma-MSH, ACTH, alpha-MSH, delta-MSH, beta-MSH, and beta-END are located at prePOMC (76-87), (120-158), (120-132), (212-227), (275-290), and (293-325), respectively. delta-MSH, originally found in elasmobranch POMCs, was also present in ratfish POMC, suggesting this structure might have appeared after the divergence of chondrichthians from the ancestral lineage. Thus, we demonstrated the common occurrence of four MSHs in chondrichthian POMC and established a clear understanding of the molecular evolution of POMC in gnathostomes.