RESUMO
BACKGROUND: The products of cyp19, dax, foxl2, mis, sf1 and sox9 have each been associated with sex-determining processes among vertebrates. We provide evidence for expression of these regulators very early in salmonid development and in tissues outside of the hypothalamic-pituitary-adrenal/gonadal (HPAG) axis. Although the function of these factors in sexual differentiation have been defined, their roles in early development before sexual fate decisions and in tissues beyond the brain or gonad are essentially unknown. RESULTS: Bacterial artificial chromosomes containing salmon dax1 and dax2, foxl2b and mis were isolated and the regulatory regions that control their expression were characterized. Transposon integrations are implicated in the shaping of the dax and foxl2 loci. Splice variants for cyp19b1 and mis in both embryonic and adult tissues were detected and characterized. We found that cyp19b1 transcripts are generated that contain 5'-untranslated regions of different lengths due to cryptic splicing of the 3'-end of intron 1. We also demonstrate that salmon mis transcripts can encode prodomain products that present different C-termini and terminate before translation of the MIS hormone. Regulatory differences in the expression of two distinct aromatases cyp19a and cyp19b1 are exerted, despite transcription of their transactivators (ie; dax1, foxl2, sf1) occurring much earlier during embryonic development. CONCLUSIONS: We report the embryonic and extragonadal expression of dax, foxl2, mis and other differentiation factors that indicate that they have functions that are more general and not restricted to steroidogenesis and gonadogenesis. Spliced cyp19b1 and mis transcripts are generated that may provide regulatory controls for tissue- or development-specific activities. Selection of cyp19b1 transcripts may be regulated by DAX-1, FOXL2 and SF-1 complexes that bind motifs in intron 1, or by signals within exon 2 that recruit splicing factors, or both. The potential translation of proteins bearing only the N-terminal MIS prodomain may modulate the functions of other TGF ß family members in different tissues. The expression patterns of dax1 early in salmon embryogenesis implicate its role as a lineage determination factor. Other roles for these factors during embryogenesis and outside the HPAG axis are discussed.
Assuntos
Regulação da Expressão Gênica no Desenvolvimento , Salmo salar/genética , Diferenciação Sexual/fisiologia , Animais , Feminino , Perfilação da Expressão Gênica , Regulação da Expressão Gênica no Desenvolvimento/genética , Regulação da Expressão Gênica no Desenvolvimento/fisiologia , Hibridização in Situ Fluorescente , Masculino , Regiões Promotoras Genéticas/genética , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Diferenciação Sexual/genéticaRESUMO
BACKGROUND: The Atlantic salmon (Salmo salar) immunoglobulin heavy chain (IgH) locus possesses two parallel IgH isoloci (IGH-A and IGH-B), that are related to the genomic duplication event in the family Salmonidae. These duplicated IgH loci in Atlantic salmon provide a unique opportunity to examine the mechanisms of genome diversity and genome evolution of the IgH loci in vertebrates. In this study, we defined the structure of these loci in Atlantic salmon, and sequenced 24 bacterial artificial chromosome (BAC) clones that were assembled into the IGH-A (1.1 Mb) and IGH-B (0.9 Mb) loci. In addition, over 7,000 cDNA clones from the IgH variable (VH) region have been sequenced and analyzed. RESULTS: The present study shows that the genomic organization of the duplicated IgH loci in Atlantic salmon differs from that in other teleosts and other vertebrates. The loci possess multiple Cτ genes upstream of the Cµ region, with three of the Cτ genes being functional. Moreover, the duplicated loci possess over 300 VH segments which could be classified into 18 families. This is the largest number of VH families currently defined in any vertebrate. There were significant structural differences between the two loci, indicating that both IGH-A and -B loci have evolved independently in the short time after the recent genome duplication approximately 60 mya. CONCLUSIONS: Our results indicate that the duplication of the IgH loci in Atlantic salmon significantly contributes to the increased diversity of the antibody repertoire, as compared with the single IgH locus in other vertebrates.
Assuntos
Evolução Molecular , Duplicação Gênica/genética , Genes de Cadeia Pesada de Imunoglobulina/genética , Loci Gênicos/genética , Salmo salar/genética , Animais , Oceano Atlântico , Perfilação da Expressão Gênica , Regulação da Expressão Gênica no Desenvolvimento , Rearranjo Gênico de Cadeia Pesada de Linfócito B/genética , Variação Genética , Regiões Constantes de Imunoglobulina/química , Regiões Constantes de Imunoglobulina/genética , Região Variável de Imunoglobulina/química , Região Variável de Imunoglobulina/genética , Dados de Sequência Molecular , Filogenia , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Alinhamento de SequênciaRESUMO
The complete TCR alpha/delta locus of Atlantic salmon (Salmo salar) has been characterized and annotated. In the 900 kb TCR alpha/delta locus, 292 Valpha/delta segments and 123 Jalpha/delta segments were identified. Of these, 128 Valpha/delta, 113 Jalpha, and a Jdelta segment appeared to be functional as they lacked frame shifts or stop codons. This represents the largest repertoire of Valpha/delta and Jalpha segments of any organism to date. The 128 functional Valpha/delta segments could be grouped into 29 subgroups based upon 70% nucleotide similarity. Expression data confirmed the usage of the diverse repertoire found at the genomic level. At least 99 Valpha, 13 Vdelta 86 Jalpha, 1 Jdelta, and 2 Ddelta segments were used in TCR alpha or delta transcription, and 652 unique genes were identified from a sample of 759 TCRalpha cDNA clones. Cumulatively, the genomic and expression data suggest that the Atlantic salmon T-cell receptor has enormous capacity to recognize a wide diversity of antigens.
Assuntos
Genes Codificadores da Cadeia alfa de Receptores de Linfócitos T/genética , Genes Codificadores da Cadeia delta de Receptores de Linfócitos T/genética , Salmo salar/genética , Alelos , Animais , Sequência de Bases , Regiões Determinantes de Complementaridade/genética , DNA Complementar/química , DNA Complementar/genética , Perfilação da Expressão Gênica , Biblioteca Gênica , Ordem dos Genes , Genes Codificadores da Cadeia beta de Receptores de Linfócitos T/genética , Genes Codificadores da Cadeia gama de Receptores de Linfócitos T/genética , Dados de Sequência Molecular , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Salmo salar/imunologia , Análise de Sequência de DNA , Homologia de Sequência do Ácido NucleicoRESUMO
Human papillomavirus (HPV) type 16 and 18 neutralizing antibody (NAb) titers were measured in 1,020 prenatal women in British Columbia aged 15 to 39. HPV 16 and 18 NAbs were detected in 183/1,020 (17.9%) and 97/1,020 (9.5%), respectively, and 39 (3.8%) had NAbs to both types. Titers were similar across age strata.