RESUMO
Root-specific cDNAs of glycine-rich protein (cucumber root glycine rich protein-1 and -2; CRGRP-1 and CRGRP-2) were cloned previously by use of an antiserum raised against whole xylem sap of Cucumis sativus. The accumulation of the corresponding mRNA at high levels was detected in the root-hair zone of cucumber tap root [Sakuta et al. (1998) Plant Cell Physiol. 39: 1330]. The RNA gel blot analysis with the CRGRP-1- and -2-specific probes revealed that the CRGRP genes expressed only in root but not at all in aboveground organs. When the localization of these mRNAs were examined by in situ hybridization, CRGRP mRNAs were found only in the parenchyma cells in the central cylinder of young lateral roots and it was most abundant in the cells that surrounded xylem vessels in the root-hair zone of the tap root. In immunoblotting of xylem sap collected from cucumber stem with an antiserum raised against CRGRP-1 that had been produced in an E. coli expression system, the antibodies, which did not cross-react with GRP1.8 of kidney bean, reacted with two proteins, whose mobilities corresponded to those of proteins deduced from the CRGRP-1 and -2 cDNAs. Immunohistochemical staining revealed that the CRGRPs accumulated specifically in the lignified walls of metaxylem vessels in the root, stem and leaf and in the lignified cell walls of perivascular fibers in cucumber stems. Immunostaining was also detected in the walls of metaxylem vessels and in the cell walls of adjacent sclerenchyma in the hypocotyl of kidney bean. These data clearly indicate that the novel glycine-rich proteins were produced in the vascular tissue of the root, transported systemically over a long distance via the xylem sap and immobilized in the walls of metaxylem vessels and sclerechyma cells in aboveground organs.
Assuntos
Cucumis sativus/genética , Regulação da Expressão Gênica de Plantas , Proteínas de Plantas/genética , Cucumis sativus/metabolismo , Fabaceae/genética , Imuno-Histoquímica , Dados de Sequência Molecular , Especificidade de Órgãos , Folhas de Planta/citologia , Folhas de Planta/metabolismo , Proteínas de Plantas/biossíntese , Proteínas de Plantas/isolamento & purificação , Raízes de Plantas/citologia , Raízes de Plantas/metabolismo , Caules de Planta/citologia , Caules de Planta/metabolismo , Plantas Medicinais , RNA Mensageiro/genética , Transcrição GênicaRESUMO
A novel cDNA encoding a major 30-kDa protein in xylem sap of cucumber Cucumis sativus (XSP30) was homologous to the B chains of galactose-binding lectins such as ricin and abrin. XSP30 gene was specifically expressed in roots, and XSP30 was immunologically detected only in the xylem sap, but not in any organs.
Assuntos
Cucumis sativus/genética , Lectinas de Plantas , Proteínas de Plantas/genética , Sequência de Aminoácidos , Sequência de Bases , Clonagem Molecular , DNA Complementar , Dados de Sequência Molecular , Raízes de Plantas/metabolismo , Homologia de Sequência de AminoácidosRESUMO
Candidate cDNAs for xylem sap proteins (XSPs) were isolated by screening of a cucumber cDNA library prepared from poly(A)+ RNA of cucumber roots with an antiserum raised against the XSPs of cucumber. The patterns of expression of corresponding transcripts and the sequences of the cDNAs were examined. Two cDNAs with a glycine-rich domain in each deduced amino acid sequence and whose transcripts were expressed predominantly in roots were selected for further analysis. Both cDNAs encoded proteins with a putative signal sequence, with similar non-glycine-rich domains at the amino terminal of the encoded polypeptides. The corresponding mRNAs accumulated concurrently with the formation of adventitious roots from cuttings of cucumber hypocotyls and the development of vascular tissues, and strong expression of the mRNAs was detected in the root-hair zone of tap roots. These data suggest that the production of the two novel glycine-rich proteins, which are putative XSPs, might be associated with the functions of the vascular tissues in roots.
Assuntos
Cucumis sativus/genética , Glicina , Proteínas de Plantas/genética , Raízes de Plantas/metabolismo , Sequência de Aminoácidos , Clonagem Molecular , DNA Complementar/genética , Expressão Gênica , Dados de Sequência Molecular , Proteínas de Plantas/imunologia , Raízes de Plantas/anatomia & histologia , Raízes de Plantas/genética , Sinais Direcionadores de Proteínas/genética , RNA Mensageiro/genética , RNA de Plantas/genética , Análise de Sequência de DNA , Distribuição TecidualRESUMO
A gene for carrot seed cysteine proteinase (CSCP; AB057371, AB057372) was cloned using PCR. The deduced amino acid sequence of CSCP had active sites for eukaryotic cysteine proteinases, putative signal sequences, and an endoplasmic reticulum targeting sequence. RNA gel blot analysis showed that CSCP transcripts appeared from dry seed, reached a maximum 3 days after imbibition (DAI), and disappeared by 5 DAI. In situ hybridization showed that CSCP transcripts accumulated in the endosperm of germinating seeds. This is the first report of the expression pattern of a cysteine proteinase gene in the endosperm of germinating dicotyledonous plants. The promoter of the CSCP gene had an endosperm motif and many other motifs also found in the promoters of endosperm-specific storage protein genes in monocotyledons. It is suggested that dicotyledons, like monocotyledons, have a temporal and spatial regulation system for endosperm-specific gene expression in germinating seeds.