Constitutive expression of apple endo-POLYGALACTURONASE1 in fruit induces early maturation, alters skin structure and accelerates softening.

During fruit ripening, polygalacturonases (PGs) are key contributors to the softening process in many species. Apple is a crisp fruit that normally exhibits only minor changes to cell walls and limited fruit softening. Here, we explore the effects of PG overexpression during fruit development using transgenic apple lines overexpressing the ripening-related endo-POLYGALACTURONASE1 gene. MdPG1-overexpressing (PGox) fruit displayed early maturation/ripening with black seeds, conversion of starch to sugars and ethylene production occurring by 80 days after pollination (DAP). PGox fruit exhibited a striking, white-skinned phenotype that was evident from 60 DAP and most likely resulted from increased air spaces and separation of cells in the hypodermis due to degradation of the middle lamellae. Irregularities in the integrity of the epidermis and cuticle were also observed. By 120 DAP, PGox fruit cracked and showed lenticel-associated russeting. Increased cuticular permeability was associated with microcracks in the cuticle around lenticels and was correlated with reduced cortical firmness at all time points and extensive post-harvest water loss from the fruit, resulting in premature shrivelling. Transcriptomic analysis suggested that early maturation was associated with upregulation of genes involved in stress responses, and overexpression of MdPG1 also altered the expression of genes involved in cell wall metabolism (e.g. ß-galactosidase, MD15G1221000) and ethylene biosynthesis (e.g. ACC synthase, MD14G1111500). The results show that upregulation of PG not only has dramatic effects on the structure of the fruit outer cell layers, indirectly affecting water status and turgor, but also has unexpected consequences for fruit development.

Metabolome and proteome changes between biofilm and planktonic phenotypes of the marine bacterium Pseudoalteromonas lipolytica TC8.

A number of bacteria adopt various lifestyles such as planktonic free-living or sessile biofilm stages. This enables their survival and development in a wide range of contrasting environments. With the aim of highlighting specific metabolic shifts between these phenotypes and to improve the overall understanding of marine bacterial adhesion, a dual metabolomics/proteomics approach was applied to planktonic and biofilm cultures of the marine bacterium Pseudoalteromonas lipolytica TC8. The liquid chromatography mass spectrometry (LC-MS) based metabolomics study indicated that membrane lipid composition was highly affected by the culture mode: phosphatidylethanolamine (PEs) derivatives were over-produced in sessile cultures while ornithine lipids (OLs) were more specifically synthesized in planktonic samples. In parallel, differences between proteomes revealed that peptidases, oxidases, transcription factors, membrane proteins and the enzymes involved in histidine biosynthesis were over-expressed in biofilms while proteins involved in heme production, nutrient assimilation, cell division and arginine/ornithine biosynthesis were specifically up-regulated in free-living cells.

Discrimination of Four Marine Biofilm-Forming Bacteria by LC-MS Metabolomics and Influence of Culture Parameters.

Most marine bacteria can form biofilms, and they are the main components of biofilms observed on marine surfaces. Biofilms constitute a widespread life strategy, as growing in such structures offers many important biological benefits. The molecular compounds expressed in biofilms and, more generally, the metabolomes of marine bacteria remain poorly studied. In this context, a nontargeted LC-MS metabolomics approach of marine biofilm-forming bacterial strains was developed. Four marine bacteria, Persicivirga (Nonlabens) mediterranea TC4 and TC7, Pseudoalteromonas lipolytica TC8, and Shewanella sp. TC11, were used as model organisms. The main objective was to search for some strain-specific bacterial metabolites and to determine how culture parameters (culture medium, growth phase, and mode of culture) may affect the cellular metabolism of each strain and thus the global interstrain metabolic discrimination. LC-MS profiling and statistical partial least-squares discriminant analyses showed that the four strains could be differentiated at the species level whatever the medium, the growth phase, or the mode of culture (planktonic vs biofilm). A MS/MS molecular network was subsequently built and allowed the identification of putative bacterial biomarkers. TC8 was discriminated by a series of ornithine lipids, while the P. mediterranea strains produced hydroxylated ornithine and glycine lipids. Among the P. mediterranea strains, TC7 extracts were distinguished by the occurrence of diamine derivatives, such as putrescine amides.

Transcriptome Responses of Ripe Cherry Tomato Fruit Exposed to Chilling and Rewarming Identify Reversible and Irreversible Gene Expression Changes.

Tomato fruit stored below 12°C lose quality and can develop chilling injury upon subsequent transfer to a shelf temperature of 20°C. The more severe symptoms of altered fruit softening, uneven ripening and susceptibility to rots can cause postharvest losses. We compared the effects of exposure to mild (10°C) and severe chilling (4°C) on the fruit quality and transcriptome of 'Angelle', a cherry-type tomato, harvested at the red ripe stage. Storage at 4°C (but not at 10°C) for 27 days plus an additional 6 days at 20°C caused accelerated softening and the development of mealiness, both of which are commonly related to cell wall metabolism. Transcriptome analysis using RNA-Seq identified a range of transcripts encoding enzymes putatively involved in cell wall disassembly whose expression was strongly down-regulated at both 10 and 4°C, suggesting that accelerated softening at 4°C was due to factors unrelated to cell wall disassembly, such as reductions in turgor. In fruit exposed to severe chilling, the reduced transcript abundances of genes related to cell wall modification were predominantly irreversible and only partially restored upon rewarming of the fruit. Within 1 day of exposure to 4°C, large increases occurred in the expression of alternative oxidase, superoxide dismutase and several glutathione S-transferases, enzymes that protect cell contents from oxidative damage. Numerous heat shock proteins and chaperonins also showed large increases in expression, with genes showing peak transcript accumulation after different times of chilling exposure. These changes in transcript abundance were not induced at 10°C, and were reversible upon transfer of the fruit from 4 to 20°C. The data show that genes involved in cell wall modification and cellular protection have differential sensitivity to chilling temperatures, and exhibit different capacities for recovery upon rewarming of the fruit.

Metabolomic and proteomic changes induced by growth inhibitory concentrations of copper in the biofilm-forming marine bacterium Pseudoalteromonas lipolytica.

Copper is an essential element for living cells but this metal is present in some marine environments at such high concentrations that it can be toxic for numerous organisms. In polluted areas, marine organisms may develop specific adaptive responses to prevent cell damage. To investigate the influence of copper on the metabolism of a single organism, a dual approach combining metabolomics and proteomics was undertaken on the biofilm-forming bacterial strain Pseudoalteromonas lipolytica TC8. In order to highlight differential adaptation according to the phenotype, the response of P. lipolytica TC8 to copper stress was studied in planktonic and biofilm culture modes under growth inhibitory copper concentrations. As expected, copper exposure led to the induction of defense and detoxification mechanisms. Specific metabolite and protein profiles were thus observed in each condition (planktonic vs. biofilm and control vs. copper-treated cultures). Copper exposure seems to induce drastic changes in the lipid composition of the bacterial cell membrane and to modulate the abundance of proteins functionally known to be involved in copper cell homeostasis in both planktonic and biofilm culture modes. Much more proteins differentially expressed after copper treatment were observed in biofilms than in planktonic cells, which could indicate a more heterogeneous response of biofilm cells to this metallic stress.