In vitro resynthesis of lichenization reveals the genetic background of symbiosis-specific fungal-algal interaction in Usnea hakonensis.

Background Symbiosis often gives organisms the ability to expand ecological niches which are inaccessible as individuals. In lichen symbiosis, mutualistic relationships between lichen-forming fungi and algae and/or cyanobacteria produce unique features that make lichens adaptive to wide range of environments. This study revealed the fungal-algal interaction specific to the symbiosis in lichen using Usnea hakonensis as a model system. Results The whole genome of U. hakonensis, the fungal partner, was sequenced by using the culture isolated from a natural lichen thallus. Isolated cultures of the fungal and the algal partners were co-cultured in vitro for three months, and the thalli were successfully resynthesized into visible protrusions. Transcriptomes of resynthesized and natural thalli (symbiotic states) were compared to that of isolated cultures (non-symbiotic state). Sets of fungal and algal genes up-regulated in both symbiotic states were identified as symbiosis-related genes. Conclusion From the predicted functions of these genes, we identified the genetic background of two main features fundamental to the symbiotic lifestyle in lichen. First is an establishment of fungal symbiotic interface: (a) production of a hydrophobic layer that ensheaths fungal and algal cells; and (b) remodeling of cell walls at fungal-algal contact sites. Second is a symbiosis-specific nutrient flow: (a) the algal supply of photosynthetic product to the fungus; and (b) the fungal supply of phosphorous and nitrogen compounds to the alga. Since both features are widespread among lichens, our result may indicate the genetic basis of lichen symbiosis.

Molecular characterization and expression pattern of sorbitol transporter gene PbSOT2 in Pear (Pyrus bretschneideri Rehd.) fruit

Sorbitol is a primary photosynthetic product and the principal photosynthetic transport substance in plants of the Rosaceae. Sorbitol transporters in the major facilitator superfamily (MFS) are important for phloem loading and sorbitol uptake into sink tissues. Here we report the cloning, localization, and expression analysis of a sorbitol transporter in fruit of Pyrus bretschneideri Rehd. cv. “Yali.” This clone, named PbSOT2, encoded a 537-aa protein with a calculated molecular mass of 57.92 kDa. The predicted protein had 12 transmembrane domains and belonged to the MFS carriers. PbSOT2 was sub-cellularly targeted to the plasma membrane. The expression of PbSOT2 was highest during the rapid enlargement phase of fruit (100 days after full bloom). In addition, the sorbitol content in fruit fluctuated within certain limits, but its proportion of total sugars decreased continuously. This work shows that PbSOT2 may play a role in fruit enlargement and the accumulation of hexose during fruit development.

Response of mannitol-producing Arabidopsis thaliana to abiotic stress

In celery, mannitol is a primary photosynthetic product that is associated with celery’s exceptional salt tolerance. Arabidopsis plants transformed with celery’s mannose-6-phosphate reductase (M6PR) gene produce mannitol and grow normally in the absence of stress. Daily analysis of the increase in growth (fresh and dry weight, leaf number, leaf area per plant and specific leaf weight) over a 12-day period showed less effect of salt (100 mm NaCl) on the M2 transformant than wild type (WT). Following a 12-day treatment of WT, M2 and M5 plants with 100 or 200 mm NaCl the total shoot fresh weight, leaf number, and leaf area were significantly greater in transformants than in WT plants. The efficiency of use of energy for photochemistry by PSII was measured daily under growth conditions. In WT plants treated with 100 mm NaCl, the PSII yield begin decreasing after 6 days with a 50% loss in yield after 12 days, indicating a severe loss in PSII efficiency; whereas, there was no effect on the transformants. Under atmospheric levels of CO2, growth with 200 mm NaCl caused an increase in the substomatal levels of CO2 in WT plants but not in transformants. It also caused a marked decrease in carboxylation efficiency under limiting levels of CO2 in WT compared with transformants. When stress was imposed and growth reduced by withholding water for 12 days, which resulted in a similar decrease in relative water content to salt-treated plants, there were no differences among the genotypes in PSII yields or growth. The results suggest mannitol, which is known to be a compatible solute and antioxidant, protects photosynthesis against salt-related damage to chloroplasts.

Photoassimilate Regulation of Sorbitol and Sucrose Metabolism in Peach Fruit

Along with sucrose, sorbitol represents the major photosynthetic product and the main form of translocated carbon in peach. The objective of the present study was to determine whether in peach fruit, sorbitol and sucrose enzyme activities are source-regulated, and more specifically modulated by sorbitol or sucrose availability. In two separate trials, peach fruit relative growth rate (RGR), enzyme activities, and carbohydrates were measured 1) at cell division stage before and after girdling of the shoot subtending the fruit; and 2) on 14 shoots with different leaf to fruit ratio (L:F) at cell division and cell expansion stages. Fruit RGR and sorbitol dehydrogenase (SDH) activity were significantly reduced by girdling, whereas sucrose synthase (SS), acid invertase (AI), and neutral invertase (NI) where equally active in girdled and control fruits on the fourth day after girdling. All major carbohydrates (sorbitol, sucrose, glucose, fructose and starch) were reduced on the fourth day after girdling. SDH activity was the only enzyme activity proportional to L:F in both fruit developmental stages. Peach fruit incubation in sorbitol for 24 hours also resulted in SDH activities higher than those of fruits incubated in buffer and similar to those of freshly extracted samples. Overall, our data provide some evidence for regulation of sorbitol metabolism, but not sucrose metabolism, by photoassimilate availability in peach fruit. In particular, sorbitol translocated to the fruit may function as a signal for modulating SDH activity.

Sorbitol Transporter Expression in Apple Sink Tissues: Implications for Fruit Sugar Accumulation and Watercore Development

In apple (Malus ×domestica Borkh.), where sorbitol is a primary photosynthetic product that is translocated throughout the plant, accumulation of sorbitol in sink cells appears to require an active carrier-mediated membrane transport step. Recent progress in isolation and characterization of genes for sorbitol transporters in sour cherry (Prunus cerasus L.) and mannitol transporters in celery (Apium graveolens L.) suggested that similar transporters may be present in apple tissues. A defect in these transporters could also explain the occurrence of the fruit disorder watercore, characterized by the accumulation of fluids and sorbitol in the apoplasmic free space. Our objectives therefore included isolation and characterization of genes for sorbitol transporters in apple tissues and comparisons of expression of transporter genes, especially in various sink tissues including watercored and non-watercored fruit tissues. We have isolated and characterized two sorbitol transporter genes, MdSOT1 and MdSOT2. Sequence analyses indicated that these are members of the major facilitator transporter superfamily that gives rise to highly hydrophobic integral membrane proteins. Heterologous expression and measurement of sorbitol uptake in yeast indicated that these are specific and with high affinities for sorbitol, with Kms for sorbitol of 1.0 and 7.8 mm for MdSOT1 and MdSOT2, respectively. Sorbitol transporter expression was evident in all sink tissues tested with the exception of watercore-affected fruit tissues. Sorbitol accumulation in apple sink tissues thus involves an apoplasmic active membrane transport step and watercore results from a defect in that process.