RESUMO
The ENF peptide family, so termed after the consensus sequence in their amino termini (Glu-Asn-Phe-), is assumed to play multiple important roles in defense reactions, growth regulation, and homeostasis of Lepidopteran insects. The paralytic peptide of Bombyx mori (BmPP) is one such peptide that is involved in the paralytic and plasmatocyte-spreading activities in the hemocyte immune reaction. The growth-blocking peptide of Pseudaletia separata (PsGBP), which is also a member of the ENF peptide family, has similar functions that can reportedly be attenuated by the growth-blocking peptide-binding protein (GBP-BP). Using the fluorescent differential display (FDD) technique, the differential expression pattern of genes in highly susceptible silkworm strain 306 were analyzed, following infection with B. mori nuclear polyhedrosis virus (BmNPV), and a differential band (G12(782)) was obtained from the hemolymph RNA pools. Using 5'-RACE with a specially designed primer based on the FDD study, a 1,401 bp cDNA clone was obtained containing a 1,311 bp open reading frame (ORF, GenBank accession number DQ306881). The deduced protein was highly homologous in primary structure to GBP-BP and was termed B. mori paralytic peptide-binding protein (PP-BP). The B. mori PP-BP gene is organized into two exons and only one intron, using bioinformatics searches.Using RT-PCR analysis, it was found that the B. mori PP-BP gene was expressed almost exclusively in the hemolymph. Real-time quantitative PCR analysis indicated that the B. mori PP-BP mRNA level in B. mori strain 306 exposed to BmNPV was much higher than that in B. mori strain without the virus infection. This result implies that the B. mori PP-BP is related to the cellular immune response after BmNPV invades the hemolymph.
Assuntos
Bombyx/genética , Bombyx/virologia , Proteínas de Insetos/genética , Neuropeptídeos/metabolismo , Nucleopoliedrovírus/fisiologia , Sequência de Aminoácidos , Animais , Sequência de Bases , Bombyx/imunologia , Clonagem Molecular , Biologia Computacional , DNA Complementar/genética , Perfilação da Expressão Gênica , Genes de Insetos/genética , Hemolinfa/metabolismo , Hemolinfa/virologia , Proteínas de Insetos/química , Proteínas de Insetos/metabolismo , Larva/genética , Larva/imunologia , Larva/virologia , Dados de Sequência Molecular , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Análise de Sequência de DNA , Fatores de TempoRESUMO
The genome of Bombyx mori densonucleosis virus type 3 (China isolate) contains two kinds different single-stranded linear DNA molecules (VD1, VD2). The VD2 was purified and cloned into the pUC119 vector, and the complete nucleotide sequence of VD2 was determined. Sequence analysis showed that the VD2 genome sequence consisted of 6022 nts, including inverted terminal repeats (ITRs) of 524 nts. In the viral genome, two major open reading frames (ORF1 and ORF2) in the plus strand and one minor ORF (ORF3) in the complementary strand were identified. Computer analysis suggested the plus stand ORF1 and the minus strand ORF3 most likely encode the major nonstructural protein, while the plus stand ORF2 most likely encodes the major structural protein. Comparing the complete genome sequence of BmDNV-3 VD2 with that of BmDNV-2 VD2 (Yamanashi isolate) demonstrated that they shared 97.7% genome sequence in the VD2 region, with substitutions of 132 nucleotides, deletions of 11 nucleotides and insertions of 2 nucleotides in the VD2 region of BmDNV-3. The results suggested that BmDNV-3 is closely related to BmDNV-2, but with some differences, giving a better understanding about the variation of the viruses and providing clues to their evolution.
Assuntos
Bombyx/virologia , Densovirus/genética , Animais , Sequência de Bases , China , Clonagem Molecular , DNA de Cadeia Simples/genética , DNA Viral/genética , Densovirus/classificação , Evolução Molecular , Genoma Viral/genética , Genômica , Dados de Sequência Molecular , Fases de Leitura Aberta , Poli A/genética , Análise de Sequência de DNA , Sequências Repetidas TerminaisRESUMO
The microsporidian Nosema antheraeae is a pathogen that infects the Chinese oak silkworm, Antheraea pernyi. We sequenced the complete small subunit (SSU) rRNA gene and the internal transcribed spacer (ITS) of N. antheraeae, and compared the SSU rRNA sequences in other microsporidia. The results indicated that Nosema species, including N. antheraeae, formed two distinct clades, consistent with previous observations. Furthermore, N. antheraeae is clustered with N. bombycis with high bootstrap support. The organization of the rRNA gene of N. antheraeae is LSU-ITS1-SSU-ITS2-5S, also following a pattern similar to the Nosema type species, N. bombycis. Thus, N. antheraeae is a Nosema species and has a close relationship to N. bombycis.