RESUMO
Two representative genes for the 54 kDa protein subunit of the signal recognition particle (SRP54) of tomato were cloned. It was shown that both genes are expressed in the tomato cv. Rentita. SRP54 is encoded by nine exons distributed over 10 kb of genomic sequence. The amino acid sequences deduced for the two SRP54 genes are 92% identical and the calculated protein size is 55 kDa. Like the homologous proteins isolated from other eukaryotes, the tomato SRP54 is evidently divided into two domains. As deduced from sequence motif identity, the N-terminally located G-domain can be assumed to have GTPase activity. The C-terminal part of the protein is methionine rich (14% methionine) and represents the M-domain. In in vitro binding experiments, SRP54 of tomato was able to attach to the 7S RNA of tomato, its natural binding partner in the SRP. This interaction can only take place in a trimeric complex consisting of 7S RNA, SRP54 and SRP19. The latter protein subunit of the SRP complex is assumed to induce a conformational change in the 7S RNA. The human SRP19 was able to mediate the binding of the tomato SRP54 to the 7S RNA, irrespective of whether this latter originated from tomato or man.
Assuntos
Proteínas de Plantas/química , Proteínas de Plantas/fisiologia , Partícula de Reconhecimento de Sinal/química , Partícula de Reconhecimento de Sinal/fisiologia , Solanum lycopersicum/química , Sequência de Aminoácidos , Sequência de Bases , Clonagem Molecular , Citoplasma/química , Humanos , Solanum lycopersicum/genética , Dados de Sequência Molecular , Conformação de Ácido Nucleico , Proteínas de Plantas/genética , Proteínas de Ligação a RNA/química , Homologia de Sequência de Aminoácidos , Partícula de Reconhecimento de Sinal/genéticaRESUMO
Two chalcone synthase genes in maize have been cloned and molecularly characterized to be the C2 and the Whp (white pollen) locus. The two genes have highly homologous exon sequences but differ considerably in sequences 5' upstream and 3' downstream of the coding region, as well as in their introns. Northern and Western experiments of chalcone synthase expression in various tissues and in different genotypes indicated that C2 and Whp are differently regulated. The expression of Whp in maize aleurone is dependent on the presence of the recessive allele of the gene intensifier (in). The regulatory effect of in on Whp expression is not detectable at the transcriptional level, but seems to take place during translation.
Assuntos
Aciltransferases/genética , Antocianinas/genética , Regulação da Expressão Gênica , Biossíntese de Proteínas , Zea mays/genética , Sequência de Aminoácidos , Sequência de Bases , Northern Blotting , Southern Blotting , Western Blotting , Clonagem Molecular , Dados de Sequência Molecular , Poli A/genética , Reação em Cadeia da Polimerase , Regiões Promotoras Genéticas , RNA/genética , RNA Mensageiro , Mapeamento por Restrição , Homologia de Sequência do Ácido Nucleico , Transcrição GênicaRESUMO
Beet necrotic yellow vein virus RNA-3 and RNA-4, produced as full-length biologically active transcripts in vitro, can undergo spontaneous internal deletions when inoculated onto Chenopodium quinoa leaves along with RNA-1 and -2. The deletion process is specific, giving rise to only a few major species, and can be rapid; deleted forms appear after only one or two passages in leaves. In one of the shortened forms of RNA-4, the deletion precisely eliminated one copy of a 15 nucleotide (nt) direct sequence repeat from the full-length prototype sequence, suggesting that 'copy-choice' switching of the replicase-template complex from one repeat to the other during RNA replication was responsible for the generation of this deletion. The deletion found in a major shortened form of RNA-3, on the other hand, did not occur near sequence repeats but began with GU and ended with AG like a nuclear intron sequence. Thus it is possible that the deleted sequence has been removed by splicing. However, two other deletions that were characterized were not associated with either of these types of sequence feature. An approximately 600 nt 5'-terminally truncated non-encapsidated form of RNA-3 was also detected in infected plant tissue. The evidence suggests that it is a subgenomic RNA derived from RNA-3.
Assuntos
Mutação , Vírus de Plantas/genética , RNA Viral/genética , Sequência de Bases , Northern Blotting , Mapeamento Cromossômico , Genes Virais , Dados de Sequência Molecular , Plantas/microbiologia , Splicing de RNA , Sequências Repetitivas de Ácido Nucleico , Transcrição GênicaRESUMO
Open reading frames identified on the four genomic RNAs of beet necrotic yellow vein virus were cloned into bacterial expression vectors and resulting cl-fusion proteins expressed in Escherichia coli were used to raise polyclonal antibodies. This set of antisera was used to show the presence of 7 of 9 predicted viral proteins in mechanically inoculated Chenopodium quinoa leaves by the Western blot technique. Viral coat protein (p22) and its readthrough protein p85 encoded by RNA-2 could be detected in all subcellular fractions. Two other RNA-2-encoded proteins, p42 and p13, are predominantly associated with membranous structures. Another RNA-2-encoded protein, p14, as well as the two polypeptides p25 and p31, encoded by RNA-3 and -4, respectively, are soluble proteins. The viral proteins could first be detected about the time lesions became visible and increased thereafter except for p85, in which case the amount of the soluble form decreased with time. No protein could be detected corresponding to the RNA-1-encoded p237 protein or to the p15 species encoded by open reading frame V of RNA-2.