Alteration of cell-wall porosity is involved in osmotic stress-induced enhancement of aluminium resistance in common bean (Phaseolus vulgaris L.)

Aluminium (Al) toxicity and drought are the two major abiotic stress factors limiting common bean production in the tropics. Using hydroponics, the short-term effects of combined Al toxicity and drought stress on root growth and Al uptake into the root apex were investigated. In the presence of Al stress, PEG 6000 (polyethylene glycol)-induced osmotic (drought) stress led to the amelioration of Al-induced inhibition of root elongation in the Al-sensitive genotype VAX 1. PEG 6000 (>> PEG 1000) treatment greatly decreased Al accumulation in the 1 cm root apices even when the roots were physically separated from the PEG solution using dialysis membrane tubes. Upon removal of PEG from the treatment solution, the root tips recovered from osmotic stress and the Al accumulation capacity was quickly restored. The PEG-induced reduction of Al accumulation was not due to a lower phytotoxic Al concentration in the treatment solution, reduced negativity of the root apoplast, or to enhanced citrate exudation. Also cell-wall (CW) material isolated from PEG-treated roots showed a low Al-binding capacity which, however, was restored after destroying the physical structure of the CW. The comparison of the Al(3+), La(3+), Sr(2+), and Rb(+) binding capacity of the intact root tips and the isolated CW revealed the specificity of the PEG 6000 effect for Al. This could be due to the higher hydrated ionic radius of Al(3+) compared with other cations (Al(3+) >> La(3+) > Sr(2+) > Rb(+)). In conclusion, the results provide circumstantial evidence that the osmotic stress-inhibited Al accumulation in root apices and thus reduced Al-induced inhibition of root elongation in the Al-sensitive genotype VAX 1 is related to the alteration of CW porosity resulting from PEG 6000-induced dehydration of the root apoplast.

Identification of superior reference genes for data normalisation of expression studies via quantitative PCR in hybrid roses (Rosa hybrida)

Background: Gene expression studies are a prerequisite for understanding the biological function of genes. Because of its high sensitivity and easy use, quantitative PCR (qPCR) has become the gold standard for gene expression quantification. To normalise qPCR measurements between samples, the most prominent technique is the use of stably expressed endogenous control genes, the so called reference genes. However, recent studies show there is no universal reference gene for all biological questions. Roses are important ornamental plants for which there has been no evaluation of useful reference genes for gene expression studies. Results: We used three different algorithms (BestKeeper, geNorm and NormFinder) to validate the expression stability of nine candidate reference genes in different rose tissues from three different genotypes of Rosa hybrida and in leaves treated with various stress factors. The candidate genes comprised the classical "housekeeping genes" (Actin, EF-1α, GAPDH, Tubulin and Ubiquitin), and genes showing stable expression in studies in Arabidopsis (PP2A, SAND, TIP and UBC). The programs identified no single gene that showed stable expression under all of the conditions tested, and the individual rankings of the genes differed between the algorithms. Nevertheless the new candidate genes, specifically, PP2A and UBC, were ranked higher as compared to the other traditional reference genes. In general, Tubulin showed the most variable expression and should be avoided as a reference gene. Conclusions: Reference genes evaluated as suitable in experiments with Arabidopsis thaliana were stably expressed in roses under various experimental conditions. In most cases, these genes outperformed conventional reference genes, such as EF1-α and Tubulin. We identified PP2A, SAND and UBC as suitable reference genes, which in different combinations may be used for normalisation in expression analyses via qPCR for different rose tissues and stress treatments. However, the vast genetic variation found within the genus Rosa, including differences in ploidy levels, might also influence expression stability of reference genes, so that future research should also consider different genotypes and ploidy levels.

Transcriptome, carbohydrate, and phytohormone analysis of Petunia hybrida reveals a complex disturbance of plant functional integrity under mild chilling stress

Cultivation of chilling-tolerant ornamental crops at lower temperature could reduce the energy demands of heated greenhouses. To provide a better understanding of how sub-optimal temperatures (12 degrees C vs. 16 degrees C) affect growth of the sensitive Petunia hybrida cultivar 'SweetSunshine Williams', the transcriptome, carbohydrate metabolism, and phytohormone homeostasis were monitored in aerial plant parts over 4 weeks by use of a microarray, enzymatic assays and GC-MS/MS. The data revealed three consecutive phases of chilling response. The first days were marked by a strong accumulation of sugars, particularly in source leaves, preferential up-regulation of genes in the same tissue and down-regulation of several genes in the shoot apex, especially those involved in the abiotic stress response. The midterm phase featured a partial normalization of carbohydrate levels and gene expression. After 3 weeks of chilling exposure, a new stabilized balance was established. Reduced hexose levels in the shoot apex, reduced ratios of sugar levels between the apex and source leaves and a higher apical sucrose/hexose ratio, associated with decreased activity and expression of cell wall invertase, indicate that prolonged chilling induced sugar accumulation in source leaves at the expense of reduced sugar transport to and reduced sucrose utilization in the shoot. This was associated with reduced levels of indole-3-acetic acid and abscisic acid in the apex and high numbers of differentially, particularly up-regulated genes, especially in the source leaves, including those regulating histones, ethylene action, transcription factors, and a jasmonate-ZIM-domain protein. Transcripts of one Jumonji C domain containing protein and one expansin accumulated in source leaves throughout the chilling period. The results reveal a dynamic and complex disturbance of plant function in response to mild chilling, opening new perspectives for the comparative analysis of differently tolerant cultivars.

Physiological and molecular analysis of the interaction between aluminium toxicity and drought stress in common bean (Phaseolus vulgaris)

Aluminium (Al) toxicity and drought are two major factors limiting common bean (Phaseolus vulgaris) production in the tropics. Short-term effects of Al toxicity and drought stress on root growth in acid, Al-toxic soil were studied, with special emphasis on Al-drought interaction in the root apex. Root elongation was inhibited by both Al and drought. Combined stresses resulted in a more severe inhibition of root elongation than either stress alone. This result was different from the alleviation of Al toxicity by osmotic stress (-0.60 MPa polyethylene glycol) in hydroponics. However, drought reduced the impact of Al on the root tip, as indicated by the reduction of Al-induced callose formation and MATE expression. Combined Al and drought stress enhanced up-regulation of ACCO expression and synthesis of zeatin riboside, reduced drought-enhanced abscisic acid (ABA) concentration, and expression of NCED involved in ABA biosynthesis and the transcription factors bZIP and MYB, thus affecting the regulation of ABA-dependent genes (SUS, PvLEA18, KS-DHN, and LTP) in root tips. The results provide circumstantial evidence that in soil, drought alleviates Al injury, but Al renders the root apex more drought-sensitive, particularly by impacting the gene regulatory network involved in ABA signal transduction and cross-talk with other phytohormones necessary for maintaining root growth under drought.