Detection and subcellular localization of dehydrin-like proteins in quinoa (Chenopodium quinoa Willd.) embryos.

The aim of this study was to characterize the dehydrin content in mature embryos of two quinoa cultivars, Sajama and Baer La Unión. Cultivar Sajama grows at 3600-4000 m altitude and is adapted to the very arid conditions characteristic of the salty soils of the Bolivian Altiplano, with less than 250 mm of annual rain and a minimum temperature of -1 degrees C. Cultivar Baer La Unión grows at sea-level regions of central Chile and is adapted to more humid conditions (800 to 1500 mm of annual rain), fertile soils, and temperatures above 5 degrees C. Western blot analysis of embryo tissues from plants growing under controlled greenhouse conditions clearly revealed the presence of several dehydrin bands (at molecular masses of approximately 30, 32, 50, and 55 kDa), which were common to both cultivars, although the amount of the 30 and 32 kDa bands differed. Nevertheless, when grains originated from their respective natural environments, three extra bands (at molecular masses of approximately 34, 38, and 40 kDa), which were hardly visible in Sajama, and another weak band (at a molecular mass of approximately 28 kDa) were evident in Baer La Unión. In situ immunolocalization microscopy detected dehydrin-like proteins in all axis and cotyledon tissues. At the subcellular level, dehydrins were detected in the plasma membrane, cytoplasm and nucleus. In the cytoplasm, dehydrins were found associated with mitochondria, rough endoplasmic reticulum cisternae, and proplastid membranes. The presence of dehydrins was also recognized in the matrix of protein bodies. In the nucleus, dehydrins were associated with the euchromatin. Upon examining dehydrin composition and subcellular localization in two quinoa cultivars belonging to highly contrasting environments, we conclude that most dehydrins detected here were constitutive components of the quinoa seed developmental program, but some of them (specially the 34, 38, and 40 kDa bands) may reflect quantitative molecular differences associated with the adaptation of both cultivars to contrasting environmental conditions.

Detection of dehydrin-like proteins in embryos and endosperm of mature Euterpe edulis seeds.

Euterpe edulis Martius, a tropical palm species characterized as highly recalcitrant, accumulated dehydrin proteins in both the endosperm and the embryo of the mature seed, as detected by Western blot analysis and immunogold electron microscopy. Three major bands at molecular masses of approximately 16, 18, and 24 kDa were identified in both samples analysed. Immunogold electron microscopy studies detected the presence of dehydrins in the embryo and endosperm. In both cases, dehydrins were immunolocalized in cytoplasm and chromatin. No labelling associated with either membranes or organelles was detected. It is known that dehydrins are produced as part of the developmental program of orthodox seeds and are also present in some recalcitrant seeds of temperate regions. The constitutive presence of dehydrins in embryos of extremely recalcitrant species of tropical origin has not been previously reported.

Sequencing of the bicistronic genome segments S7 and S9 of Mal de Río Cuarto virus (Fijivirus, Reoviridae) completes the genome of this virus.

The nucleotide sequences of genomic segments S7 and S9 of Mal de Río Cuarto virus (MRCV, Fijivirus group II) have been determined, thus completing the entire genome sequence of the virus. These segments showed a non-overlapping bicistronic structure, as in other members of the genus. MRCV S7 ORF-1 had a length of 1086 bp and encoded a 41.5 kDa putative polypeptide, whereas MRCV S7 ORF-2 had a length of 930 bp and encoded a 36.8 kDa putative polypeptide. Proteins of 39 and 20.5 kDa were predicted for the 1014 bp long MRCV S9 ORF-1 and the 537 bp long MRCV S9 ORF-2, respectively. The terminal 5' and 3' sequences of both segments were 5'AAGUUUUU3' and 5'CAGCUnnnGUC3', respectively. Specific imperfect inverted repeats of each segment were identified. Comparison of the predicted proteins with those of related virus genome segments counterparts in maize rough dwarf virus (MRDV) and rice black streaked dwarf virus (RBSDV), showed 64.5-44.3% identities. These values are lower than those resulting from comparisons between MRDV and RBSDV. The topology of the trees obtained using the complete nucleotide and amino acid sequences of MRCV S7 and MRCV S9 was consistent with the analysis of the other MRCV segments previously published.

Sequence analysis of genome segments S5 and S10 of Mal de Rio Cuarto virus (Fijivirus, Reoviridae).

Mal de Rio Cuarto virus (MRCV) was recently described as a new species of the genus Fijivirus, family Reoviridae. The nucleotide sequence of two MRCV genome segments was determined. MRCV S5 and S10 were predicted to encode proteins of 106.9 and 63.5 kDa respectively. The protein coded by MRCV S5 had 62.8% and 35.7% identity to fijiviruses RBSDV S5 and FDV S5 coded proteins, and contained a rarely reported type-1 C-terminal peroxisomal targeting signal. The protein coded by MRCV S10 had identity levels of 72.4% and 21.7% to the major outer capsid proteins of fijiviruses RBSDV S10 and NLRV S8.

Sequence analysis of genome segments S4 and S8 of Mal de Río Cuarto virus (MRCV): evidence that the virus should be a separate Fijivirus species.

This is the first sequence-based characterization of Mal de Río Cuarto virus (MRCV), currently classified as a variant of Maize rough dwarf virus (MRDV) and exclusively found in South America. We sequenced and analyzed genome segments S4 and S8. MRCV S4 coded for a putative 131.67 kDa protein while MRCV S8 coded for a putative 68.26 kDa protein containing an ATP/GTP-binding motif. The 5' and 3' ends of MRCV segments, were 5'AAGUUUUU3' and 5'CAGCUnnnGUC3', respectively. Prediction of secondary structure of both segments coding strands showed that terminal regions were able to form structures that are proposed to be replication and packaging signals. MRCV S4 showed identity to members of Fijivirus as well as to two other genera of the Reoviridae family. MRCV S8 revealed identity with Rice black streaked dwarf virus (RBSDV) S8, MRDV S7, Oat sterile dwarf virus (OSDV) S9 and Nilaparvata lugens reovirus (NLRV) S7. While MRDV and RBSDV segments are highly homologous between each other, MRCV identity levels with them was considerably lower. We discussed the evolutionary relationships of MRCV to other Reoviridae, and based on phylogenetic analysis we proposed that although MRCV is related to MRDV, it could be regarded as a new species of the Fijivirus genus.