Mint essential oil can induce or inhibit potato sprouting by differential alteration of apical meristem

Sprouting of potatoes during storage, due to tuber dormancy release, is associated with weight loss and softening. Sprout-preventing chemicals, such as chlorpropham (CIPC), can negatively impact the environment and human health. Monthly thermal fogging with mint (Mentha spicata L.) essential oil (MEO) inhibited sprouting in eight potato cultivars during large-volume 6-month storage: the tubers remained firm with 38% lower weight loss after 140 days of storage. The sprout-inhibitory action may be nullified: treated tubers washed with water resumed sprouting within days, with reduced apical dominance. MEO application caused local necrosis of the bud meristem, and a few weeks later, axillary bud (AX) growth was induced in the same sprouting eye. MEO components analysis showed that 73% of its content is the monoterpene R-carvone. Tubers treated with synthetic R-carvone in equivalent dose, 4.5 μl l⁻¹, showed an inhibitory effect similar to that of MEO. Surprisingly, 0.5 μl l⁻¹ of MEO or synthetic R-carvone catalyzed AX sprouting in the tuber. To the best of our knowledge, this is the first report of an essential oil vapor inducing early sprouting of potato tubers. R-carvone caused visible damage to the meristem membrane at sprout-inhibiting, but not sprout-inducing doses, suggesting different underlying mechanisms. After 5 days’ exposure to R-carvone, its derivatives transcarveol and neo-dihydrocarveol were found in buds of tubers treated with the inhibitory dose, suggesting biodegradation. These experiments demonstrate the potential of MEO vapor as an environmentally friendly alternative to CIPC in stored potatoes and as a research tool for the control of sprouting in plants.     

Proteorhodopsin-Bearing Bacteria in Antarctic Sea Ice

Proteorhodopsins (PRs) are widespread bacterial integral membrane proteins that function as light-driven proton pumps. Antarctic sea ice supports a complex community of autotrophic algae, heterotrophic bacteria, viruses, and protists that are an important food source for higher trophic levels in ice-covered regions of the Southern Ocean. Here, we present the first report of PR-bearing bacteria, both dormant and active, in Antarctic sea ice from a series of sites in the Ross Sea using gene-specific primers. Positive PR sequences were generated from genomic DNA at all depths in sea ice, and these sequences aligned with the classes Alphaproteobacteria, Gammaproteobacteria, and Flavobacteria. The sequences showed some similarity to previously reported PR sequences, although most of the sequences were generally distinct. Positive PR sequences were also observed from cDNA reverse transcribed from RNA isolated from sea ice samples. This finding indicates that these sequences were generated from metabolically active cells and suggests that the PR gene is functional within sea ice. Both blue-absorbing and green-absorbing forms of PRs were detected, and only a limited number of blue-absorbing forms were found and were in the midsection of the sea ice profile in this study. Questions still remain regarding the protein''s ecological functions, and ultimately, field experiments will be needed to establish the ecological and functional role of PRs in the sea ice ecosystem.Proteorhodopsins (PRs) are retinal binding bacterial integral membrane proteins that function as light-driven proton pumps (9, 10) and belong to the microbial rhodopsin superfamily of proteins (54). Since the first reported PR sequence from members of SAR86 clade marine (class Gammaproteobacteria) in 2000 (9), many other PR-bearing bacteria have been identified in a range of marine habitats (5, 18, 20, 24, 25, 46, 62). In the recent Global Ocean Sampling (GOS) expedition, almost 4,000 PR sequences from 41 distinct surface marine environments were acquired, demonstrating that these PR genes are extremely abundant in the genomes of ocean bacterioplankton (46). In fact, PR-containing bacteria account for 13% of the community in the Mediterranean Sea and Red Sea and 70% of the community in the Sargasso Sea (18, 46, 49, 60). These light-harvesting bacteria are present in three major marine classes of bacteria: the Alphaproteobacteria, Gammaproteobacteria, and Flavobacteria. In addition, two distinct PR genes encode pigments with “blue-absorbing” and “green-absorbing” properties, which is achieved by a substitution at a single amino acid position, which thereby functions as a spectral tuning switch (10, 37, 48).Sea ice represents a complex physicochemical environment in polar regions and covers up to 13% of the Earth''s surface (59). Although extreme gradients of temperature, salinity, nutrient availability, and light stratify the ice matrix from the surface to the ice-water interface (41), the sea ice habitat nevertheless supports a diverse microbial community of phytoplankton, Bacteria, Archaea, viruses, and protists that grow in liquid brine channels within the ice (14, 35, 56). This sea ice microbial community (SIMCO) is highly metabolically active despite being unable to avoid the extreme environmental conditions that they experience (39). In fact, very-high-standing stocks of the SIMCO exist in many regions of the Southern Ocean. For example, the concentration of chlorophyll a, a proxy for microalgal biomass, typically reaches 200 mg m⁻² in the Ross Sea, while the concentration of chlorophyll a in the water column below is approximately 2 orders of magnitude less (47), and the percentage of metabolically active bacteria (32% [39]) is significantly higher than the 10% observed for temperate marine systems (36). The SIMCO is thus a major source of biomass in ice-covered regions of the Southern Ocean (59), providing a critical food source for grazing zooplankton (and, consequently, also for higher trophic levels) for much of the year (3, 59). This biomass is of particular importance during the darkness of the polar winter, where the bottom-ice community is the only available food source for juvenile krill. These grazers absolutely rely on the sea ice microbial community to survive, as the water lacks other food sources (6, 28).In the past decade, reports of the widespread occurrence of bacteriochlorophyll and PR pigments in planktonic marine bacteria have challenged the assumption that chlorophyll a is the only principal light-capturing pigment in ocean surface waters. These alternative pigments may in fact play a critical role in light energy harvesting for microbial metabolism in various aquatic ecosystems (5, 10, 25, 40, 49). It has been proposed that energy, rather than nutrient conservation, is important for the regulation of productivity (7). PR-containing phototrophic eubacteria could play a significant role in the energy budget of cells in the photic zone in marine environments (15). PR sequences have been detected in the Southern Ocean (9), but to our knowledge, there have been no reports of PR-bearing bacteria within the sea ice matrix.The majority of the microbial rhodopsin genes found in oceanic samples have been detected by environmental sequencing (30, 46, 48, 60). We have used degenerate PR gene primers (5) in this study to positively identify PR-bearing operational taxonomic units (OTUs) from sea ice. Also, specific bacterial mRNA can now be detected from extracted nucleic acids and used to examine gene expression and, thus, infer metabolic activity (8). With this in mind, we have generated cDNA from RNA extracted from sea ice samples. From these observations, we deduce that PR-bearing bacteria are present in sea ice and may be actively contributing to the ecosystem within this extreme microenvironment.     

Production and purification of lentiviral vectors

Lentiviral vectors offer unique versatility and robustness as vehicles for gene delivery. They can transduce a wide range of cell types and integrate into the host genome in both dividing and post-mitotic cells, resulting in long-term expression of the transgene both in vitro and in vivo. This protocol describes how lentiviral vectors can be produced, purified and titrated. High titer suspensions can be routinely prepared with relative ease: a low-titer (10(6) viral particles/ml) unpurified preparation can be obtained 3 d after transfecting cells with lentiviral vector and packaging plasmids; a high-titer (10(9) viral particles/ml) purified preparation requires 2 more days.     

Growth modulation of transgenic potato plants by heterologous expression of bacterial carbohydrate-binding module

Transgenic potato plants (Solanum tuberosum cv. Desiree) expressing the bacterial carbohydrate-binding module (CBM) family III, which is part of the Clostridium cellulovorans CBPA, under control of the CaMV 35S promoter were employed to investigate the influence of this protein on plant development. Eleven independent transgenic plants were found to express the cbm gene, at levels varying from one to four copies. Relative to non-transgenic controls, CBM-expressing plants were characterized by significantly more rapid elongation of the main stem. In addition, under both greenhouse and field conditions, the emergence rate of these plants was higher than in the controls, and their leaf area at early stages of development was larger, resulting in faster accumulation of fresh and dry weight than in control plants. Determination of cell size indicated that epidermal cells in young tissue were significantly larger in CBM-expressing than in control potato plants. These findings suggest that the CBM influence at the cellular level my cause significant alterations in plant growth both in tissue culture and in vivo under field conditions.     

The STE20/germinal center kinase POD6 interacts with the NDR kinase COT1 and is involved in polar tip extension in Neurospora crassa

Members of the Ste20 and NDR protein kinase families are important for normal cell differentiation and morphogenesis in various organisms. We characterized POD6 (NCU02537.2), a novel member of the GCK family of Ste20 kinases that is essential for hyphal tip extension and coordinated branch formation in the filamentous fungus Neurospora crassa. pod-6 and the NDR kinase mutant cot-1 exhibit indistinguishable growth defects, characterized by cessation of cell elongation, hyperbranching, and altered cell-wall composition. We suggest that POD6 and COT1 act in the same genetic pathway, based on the fact that both pod-6 and cot-1 can be suppressed by 1) environmental stresses, 2) altering protein kinase A activity, and 3) common extragenic suppressors (ropy, as well as gul-1, which is characterized here as the ortholog of the budding and fission yeasts SSD1 and Sts5, respectively). Unlinked noncomplementation of cot-1/pod-6 alleles indicates a potential physical interaction between the two kinases, which is further supported by coimmunoprecipitation analyses, partial colocalization of both proteins in wild-type cells, and their common mislocalization in dynein/kinesin mutants. We conclude that POD6 acts together with COT1 and is essential for polar cell extension in a kinesin/dynein-dependent manner in N. crassa.     

Influence of lipophilicity on the interactions of hydroxy stilbenes with cytochrome P450 3A4

Resveratrol, a polyphenol found in red wine, was recently suggested to act as an irreversible, mechanism-based inactivator of cytochrome P450 3A4 (CYP3A4). We found a significant inhibition of human CYP3A4-dependent transformation of cyclosporine by resveratrol, with IC50 = 4.5 microM. We studied the kinetics parameters of CYP3A4 transformation of resveratrol and structurally related, naturally occurring stilbenes. Resveratrol, piceid, resveratroloside, 5,4'-dihydroxy-3-O-methoxystilbene, and 5,3-dihydroxy-4'-O-methoxystilbene were all shown to inhibit hydroxylation of testosterone by CYP3A4. Both methoxy-stilbenes had lower IC50 values, ranging from 0.43 to 0.47 microM, suggesting that lipophilicity rather than number or positions of free hydroxyls (3,5 or 5,4') determines the CYP3A4 inhibition capacity of polyphenols. In line with these findings, both glucosyl-stilbenes were found to be weak inhibitors of CYP3A4. The affinity of the enzyme towards methoxy-stilbenes, expressed as apparent Km, was indeed higher than those for the parent resveratrol and its glucosides, in CYP3A4 reaction mixtures. Vmax values were similar, except for piceid. These results support the role of lipophilicity in the interaction of polyphenols with CYP3A4. It is suggested that selective structural modifications of substrates add significantly to knowledge acquired through molecular modifications of the enzyme.     

Isolation and characterization of<Emphasis Type="Italic"> Erythrobacter</Emphasis> sp. strains from the upper ocean

Seven strains of marine aerobic anoxygenic phototrophs belonging to the genus Erythrobacter were isolated. The strains were characterized regarding their physiological and biochemical properties, 16S rDNA and pufM gene sequences, morphological features, substrate preference, as well as pigment and lipid composition. All strains had functional type-2 reaction centers containing bacteriochlorophyll, served by small, light-harvesting complex 1, and were photosynthetically competent. In addition, large pools of carotenoids were found, but only some of the accessory pigments transfer energy to the reaction centers. All of the isolates were facultative photoheterotrophs. They required an organic carbon substrate for growth; however, they are able to supplement a significant fraction of their metabolic requirements with photosynthetically derived energy.Abbreviations BChl Bacteriochlorophyll - Chl Chlorophyll - Erb. Erythrobacter - Erm. Erythromicrobium - FAMEs Fatty acid methyl esters - IRFRR Infrared fast repetition rate - LH1, LH2 Light-harvesting complex 1 and 2, respectively - Por. Porphyrobacter - PUFAs Polyunsaturated fatty acids - Rsb. Roseobacter - RubisCO Ribulose-1,5-bisphosphate carboxylase/oxygenase - µ Growth rate - ₄₇₀ Functional cross-section of the photosynthetic unit at 470 nm     

Abnormal `wrinkled' cell walls and retarded development of transgenic Arabidopsis thaliana plants expressing endo-1,4-β-glucanase (cell) antisense

Transgenic Arabidopsis thaliana plants expressing cell antisense exhibit reduced levels of cell mRNA and protein compared with wild-type plants. The former display significant alterations in their phenotype. cell antisense plants have shorter stems and roots and are mechanically weaker than their wild-type counterparts. In cell antisense plants, the cell wall structure is markedly disrupted: both fluorescent confocal microscopy and scanning electron microscopy revealed `wrinkled' cell walls, thus indicating that CEL1 plays an important role in cell wall relaxation during cell growth and expansion. In cell antisense plants, the number of xylem elements per bundle is smaller than in the wild-type. In addition, both xylem elements and interfascicular fibers are significantly less lignified in the former. It is suggested that in A. thaliana, abnormal cell wall deposition affected by CEL1 depletion is associated not only with cell growth, but also with the differentiation process in the vascular and supporting tissues.Equal contributors