Water stress induces up-regulation of DOF1 and MIF1 transcription factors and down-regulation of proteins involved in secondary metabolism in amaranth roots (Amaranthus hypochondriacus L.).

Roots are the primary sites of water stress perception in plants. The aim of this work was to study differential expression of proteins and transcripts in amaranth roots (Amaranthus hypochondriacus L.) when the plants were grown under drought stress. Changes in protein abundance within the roots were examined using two-dimensional electrophoresis and LC/ESI-MS/MS, and the differential expression of transcripts was evaluated with suppression subtractive hybridisation (SSH). Induction of drought stress decreased relative water content in leaves and increased solutes such as proline and total soluble sugars in roots. Differentially expressed proteins such as SOD(Cu-Zn) , heat shock proteins, signalling-related and glycine-rich proteins were identified. Up-regulated transcripts were those related to defence, stress, signalling (Ser, Tyr-kinases and phosphatases) and water transport (aquaporins and nodulins). More noteworthy was identification of the transcription factors DOF1, which has been related to several plant-specific biological processes, and MIF1, whose constitutive expression has been related to root growth reduction and dwarfism. The down-regulated genes/proteins identified were related to cell differentiation (WOX5A) and secondary metabolism (caffeic acid O-methyltransferase, isoflavone reductase-like protein and two different S-adenosylmethionine synthetases). Amaranth root response to drought stress appears to involve a coordinated response of osmolyte accumulation, up-regulation of proteins that control damage from reactive oxygen species, up-regulation of a family of heat shock proteins that stabilise other proteins and up-regulation of transcription factors related to plant growth control.

Tryptic amaranth glutelin digests induce endothelial nitric oxide production through inhibition of ACE: antihypertensive role of amaranth peptides.

Amaranth seed proteins have a better balance of essential amino acids than cereals and legumes. In addition, the tryptic hydrolysis of amaranth proteins generates, among other peptides, angiotensin converting enzyme (ACE) inhibitory (ACEi) peptides. ACE converts angiotensin I (Ang I) into Ang II, but is also responsible for the degradation of bradykinin (BK). In contrast to Ang II, BK stimulates vasodilation modulated through endothelial nitric oxide (NO) production. The aim of the present study was to characterize the ACEi activity of amaranth trypsin-digested glutelins (TDGs) and their ability to induce endothelial NO production. An IC(50) value of 200microgml(-1) was measured for TDG inhibition of ACE. TDGs stimulated endothelial NO production in coronary endothelial cells (CEC) by 52% compared to control. The effects of TDGs were comparable to those of BK and Captopril, both used as positive controls of NO production. Consistent with these effects, TDGs induced, in a dose-dependent manner, endothelial NO-dependent vasodilation in isolated rat aortic rings. These results suggest that TDGs induce endothelial NO production and consequent vasodilation through their ACEi activity. Amaranth TDGs have a high potential as a nutraceutical food in prevention of cardiovascular diseases. Further molecular, cellular and physiological studies are currently under way and the results may contribute to a better understanding and control of cardiovascular disorders.

Globulin 11S and its mixture with L-dipalmitoylphosphatidylcholine at the air/liquid interface.

Langmuir films of globulin 11S protein, l-dipalmitoylphosphatidylcholine (L-DPPC), and mixtures of both on water and on buffer subphases were studied. Brewster angle microscopy (BAM) was used to characterize in situ the films morphology along Pi-A isotherms at the air/liquid interface. The L-DPPC monolayer on water behaved as has been reported extensively in the literature but a slight increase on surface pressure and a notable change in domain morphology is observed on buffer. This difference in domain behavior is due to the stabilization interaction of the LE phase by the buffer ions. On the other hand, the protein monolayer was prepared by direct deposit or injection below the surface. Both methods formed mostly a condensed film, with a multilayer formed by globular aggregates in the first method with the two subphases. However, the second method showed different behavior of the protein films depending on the subphase; on water the protein formed a homogeneous film with some globule aggregates, but on buffer a remarkably well-organized monolayer was observed by atomic force microscopy (AFM). Mixtures of globulin 11S and L-DPPC were prepared using both methods for the protein film formation at the air/fluid interface. BAM showed that the mixtures formed coexistence regions between two condensed phases, whose domains of both phases behave like liquids. Fingering phenomena were observed at the interface between protein-rich and L-DPPC-rich domains, which indicates that both phases are fluid. AFM images of the mixtures show the formation of protein- or L-DPPC-rich domains. The liquidlike behavior could be explained due to different sizes of the protein and the L-DPPC, the minority compound in each kind of domain produces defects making them behave as liquids. Interestingly enough, as the monolayer is compressed to higher surface pressure, the lipid molecules are squeezed out and complete separation of the protein and L-DPPC is produced. Furthermore, we present evidence that the protein/L-DPPC mixtures produce films with holes, which might indicate its tendency to form hollow aggregates that could have some relevance in water-channel formation for in vivo seed germination.

Bioactive peptides in amaranth (Amaranthus hypochondriacus) seed.

Amaranth seeds are rich in protein with a high nutritional value, but little is known about their bioactive compounds that could benefit health. The objectives of this research were to investigate the presence, characterization, and the anticarcinogenic properties of the peptide lunasin in amaranth seeds. Furthermore, to predict and identify other peptides in amaranth seed with potential biological activities. ELISA showed an average concentration of 11.1 microg lunasin equivalent/g total extracted protein in four genotypes of mature amaranth seeds. Glutelin fraction had the highest lunasin concentration (3.0 microg/g). Lunasin was also identified in albumin, prolamin and globulin amaranth protein fractions and even in popped amaranth seeds. Western blot analysis revealed a band at 18.5 kDa, and MALDI-TOF analysis showed that this peptide matched more than 60% of the soybean lunasin peptide sequence. Glutelin extracts digested with trypsin, showed the induction of apoptosis against HeLa cells. Prediction of other bioactive peptides in amaranth globulins and glutelins were mainly antihypertensive. This is the first study that reports the presence of a lunasin-like peptide and other potentially bioactive peptides in amaranth protein fractions.