Wayne L. Morris

The protein kinase C (PKC1) pathway is essential for maintaining cell integrity in yeast. Here it is shown that various forms of cell wall damage result in activation of the downstream MAP kinase Slt2/Mpk1. Several cell wall mutants displayed enhanced FKS2-lacZ expression, a known output of Slt2 activation. A similar response was obtained with wild-type cells grown in the presence of the cell wall perturbants Calcofluor white and Zymolyase. Upregulation of FKS2-lacZ in response to sublethal concentrations of these agents fully depended on the presence of Slt2. The same cell wall stress conditions resulted in dual threonine and tyrosine phosphorylation of Slt2. Both Slt2 phosphorylation and FKS2-lacZ induction could be largely prevented by providing osmotic support to the plasma membrane. Interestingly, Slt2 phosphorylation in response to cell wall damage required the putative plasma-membrane-located sensor Mid2 but not Hcs77/Wsc1. Finally, cell wall perturbation gave rise to cells with increased resistance to glucanase digestion and heat shock. These responses depended on the presence of Slt2. These results indicate that weakening of the cell wall activates the Slt2/Mpk1 MAP kinase pathway and results in compensatory changes in the cell wall.

Although significant work has been undertaken regarding the response of model and crop plants to heat shock during the acclimatory phase, few studies have examined the steady-state response to the mild heat stress encountered in temperate agriculture. In the present work, we therefore exposed tuberizing potato plants to mildly elevated temperatures (30/20 °C, day/night) for up to 5 weeks and compared tuber yield, physiological and biochemical responses, and leaf and tuber metabolomes and transcriptomes with plants grown under optimal conditions (22/16 °C). Growth at elevated temperature reduced tuber yield despite an increase in net foliar photosynthesis. This was associated with major shifts in leaf and tuber metabolite profiles, a significant decrease in leaf glutathione redox state and decreased starch synthesis in tubers. Furthermore, growth at elevated temperature had a profound impact on leaf and tuber transcript expression with large numbers of transcripts displaying a rhythmic oscillation at the higher growth temperature. RT-PCR revealed perturbation in the expression of circadian clock transcripts including StSP6A, previously identified as a tuberization signal. Our data indicate that potato plants grown at moderately elevated temperatures do not exhibit classic symptoms of abiotic stress but that tuber development responds via a diversity of biochemical and molecular signals.

Germplasm of Solanum tuberosum and Solanum phureja exhibit a wide (over 20-fold) variation in tuber carotenoid content. The levels of carotenoids during tuber development and storage were compared in a high carotenoid-accumulating S. phureja accession (DB375\1) with two S. tuberosum cultivars (Pentland Javelin and Desiree) that accumulate lower levels of tuber carotenoid. On a dry weight basis, total carotenoid levels were at a maximum early in tuber development. However, in the S. phureja accession, carotenoid levels remained at a high level throughout tuber development, whereas in the S. tuberosum accessions, carotenoid content decreased as dry weight increased. The carotenoid profiles of tissues during tuber development were analysed in greater detail by reverse phase HPLC. In S. phureja tubers at maturity the major carotenoids were zeaxanthin, antheraxanthin, and violaxanthin. Following 9 months storage at 4 degrees C the levels of zeaxanthin and antheraxanthin decreased, whereas the level of lutein increased; overall, however, there was only a small decrease in total carotenoid content. In order to explore the reasons for the wide variation in tuber carotenoid content, the expression patterns of the major genes encoding the enzymes of the carotenoid biosynthetic pathway were compared. Significant differences in the profiles were detected, suggesting that transcriptional control or mRNA stability gives rise to the large differences in tuber carotenoid content. In particular, there was an inverse trend between the level of zeaxanthin epoxidase transcript level and tuber carotenoid content in a range of potato germplasm, giving rise to an hypothesis for the regulation of carotenogenesis in potato tubers.

The factors that regulate storage organ carotenoid content remain to be fully elucidated, despite the nutritional and economic importance of this class of compound. Recent findings suggest that carotenoid pool size is determined, at least in part, by the activity of carotenoid cleavage dioxygenases. The aim of this study was to investigate whether Carotenoid Cleavage Dioxygenase4 (CCD4) activity affects potato (Solanum tuberosum) tuber carotenoid content. Microarray analysis revealed elevated expression of the potato CCD4 gene in mature tubers from white-fleshed cultivars compared with higher carotenoid yellow-fleshed tubers. The expression level of the potato CCD4 gene was down-regulated using an RNA interference (RNAi) approach in stable transgenic lines. Down-regulation in tubers resulted in an increased carotenoid content, 2- to 5-fold higher than in control plants. The increase in carotenoid content was mainly due to elevated violaxanthin content, implying that this carotenoid may act as the in vivo substrate. Although transcript level was also reduced in plant organs other than tubers, such as leaves, stems, and roots , there was no change in carotenoid content in these organs. However, carotenoid levels were elevated in flower petals from RNAi lines. As well as changes in tuber carotenoid content, tubers from RNAi lines exhibited phenotypes such as heat sprouting, formation of chain tubers, and an elongated shape. These results suggest that the product of the CCD4 reaction may be an important factor in tuber heat responses.

Potato production is often constrained by abiotic stresses such as drought and high temperatures which are often present in combination. In the present work, we aimed to identify key mechanisms and processes underlying single and combined abiotic stress tolerance by comparative analysis of tolerant and susceptible cultivars. Physiological data indicated that the cultivars Desiree and Unica were stress tolerant while Agria and Russett Burbank were stress susceptible. Abiotic stress caused a greater reduction of photosynthetic carbon assimilation in the susceptible cultivars which was associated with a lower leaf transpiration rate. Oxidative stress, as estimated by the accumulation of malondialdehyde was not induced by stress treatments in any of the genotypes with the exception of drought stress in Russett Burbank. Stress treatment resulted in increases in ascorbate peroxidase activity in all cultivars except Agria which increased catalase activity in response to stress. Transcript profiling highlighted a decrease in the abundance of transcripts encoding proteins associated with PSII light harvesting complex in stress tolerant cultivars. Furthermore, stress tolerant cultivars accumulated fewer transcripts encoding a type-1 metacaspase implicated in programmed cell death. Stress tolerant cultivars exhibited stronger expression of genes associated with plant growth and development, hormone metabolism and primary and secondary metabolism than stress susceptible cultivars. Metabolite profiling revealed accumulation of proline in all genotypes following drought stress that was partially suppressed in combined heat and drought. On the contrary, the sugar alcohols inositol and mannitol were strongly accumulated under heat and combined heat and drought stress while galactinol was most strongly accumulated under drought. Combined heat and drought also resulted in the accumulation of valine, isoleucine and lysine in all genotypes. These data indicate that single and multiple abiotic stress tolerance in potato is associated with a maintenance of CO2 assimilation and protection of PSII by a reduction of light harvesting capacity. The data further suggests that stress tolerant cultivars suppress cell death and maintain growth and development via fine tuning of hormone signaling, and primary and secondary metabolism. This study highlights potential targets for the development of stress tolerant potato cultivars.