Identification and genetic characterization of a gibberellin 2-oxidase gene that controls tree stature and reproductive growth in plum

Several dwarf plum genotypes (Prunus salicina L.), due to deficiency of unknown gibberellin (GA) signalling, were identified. A cDNA encoding GA 2-oxidase (PslGA2ox), the major gibberellin catabolic enzyme in plants, was cloned and used to screen the GA-deficient hybrids. This resulted in the identification of a dwarf plum hybrid, designated as DGO24, that exhibits a markedly elevated PslGA2ox signal. Grafting ‘Early Golden’ (EG), a commercial plum cultivar, on DGO24 (EG/D) enhanced PslGA2ox accumulation in the scion part and generated trees of compact stature. Assessment of active GAs in such trees revealed that DGO24 and EG/D accumulated relatively much lower quantities of main bioactive GAs (GA1 and GA4) than control trees (EG/M). Moreover, the physiological function of PslGA2ox was studied by determining the molecular and developmental consequences due to ectopic expression in Arabidopsis. Among several lines, two groups of homozygous transgenics that exhibited contrasting phenotypes were identified. Group-1 displayed a dwarf growth pattern typical of mutants with a GA deficiency including smaller leaves, shorter stems, and delay in the development of reproductive events. In contrast, Group-2 exhibited a ‘GA overdose’ phenotype as all the plants showed elongated growth, a typical response to GA application, even under limited GA conditions, potentially due to co-suppression of closely related Arabidopsis homologous. The studies reveal the possibility of utilizing PslGA2ox as a marker for developing size-controlling rootstocks in Prunus

Expression of auxin-binding protein1 during plum fruit ontogeny supports the potential role of auxin in initiating and enhancing climacteric ripening

Auxin-binding protein1 (ABP1) is an active element involved in auxin signaling and plays critical roles in auxin-mediated plant development. Here, we report the isolation and characterization of a putative sequence from Prunus salicina L., designated PslABP1. The expected protein exhibits a similar molecular structure to that of well-characterized maize-ABP1; however, PslABP1 displays more sequence polarity in the active-binding site due to substitution of some crucial amino-acid residues predicted to be involved in auxin-binding. Further, PslABP1 expression was assessed throughout fruit ontogeny to determine its role in fruit development. Comparing the expression data with the physiological aspects that characterize fruit-development stages indicates that PslABP1 up-regulation is usually associated with the signature events that are triggered in an auxin-dependent manner such as floral induction, fruit initiation, embryogenesis, and cell division and elongation. However, the diversity in PslABP1 expression profile during the ripening process of early and late plum cultivars seems to be due to the variability of endogenous auxin levels among the two cultivars, which consequently can change the levels of autocatalytic ethylene available for the fruit to co-ordinate ripening. The effect of auxin on stimulating ethylene production and in regulating PslABP1 was investigated. Our data suggest that auxin is involved in the transition of the mature green fruit into the ripening phase and in enhancing the ripening process in both auxin- and ethylene-dependent manners thereafter

Two highly divergent alcohol dehydrogenases of melon exhibit fruit ripening-specific expression and distinct biochemical characteristics

Alcohol dehydrogenases (ADH) participate in the biosynthetic pathway of aroma volatiles in fruit by interconverting aldehydes to alcohols and providing substrates for the formation of esters. Two highly divergent ADH genes (15% identity at the amino acid level) of Cantaloupe Charentais melon (Cucumis melo var. Cantalupensis) have been isolated. Cm-ADH1 belongs to the medium-chain zinc-binding type of ADHs and is highly similar to all ADH genes expressed in fruit isolated so far. Cm-ADH2 belongs to the short-chain type of ADHs. The two encoded proteins are enzymatically active upon expression in yeast. Cm-ADH1 has strong preference for NAPDH as a co-factor, whereas Cm-ADH2 preferentially uses NADH. Both Cm-ADH proteins are much more active as reductases with Kms 10–20 times lower for the conversion of aldehydes to alcohols than for the dehydrogenation of alcohols to aldehydes. They both show strong preference for aliphatic aldehydes but Cm-ADH1 is capable of reducing branched aldehydes such as 3-methylbutyraldehyde, whereas Cm-ADH2 cannot. Both Cm-ADH genes are expressed specifically in fruit and up-regulated during ripening. Gene expression as well as total ADH activity are strongly inhibited in antisense ACC oxidase melons and in melon fruit treated with the ethylene antagonist 1-methylcyclopropene (1-MCP), indicating a positive regulation by ethylene. These data suggest that each of the Cm-ADH protein plays a specific role in the regulation of aroma biosynthesis in melon fruit

The Slab Puzzle of the Alpine‐Mediterranean Region: Insights from a new, High‐Resolution, Shear‐Wave Velocity Model of the Upper Mantle

Mediterranean tectonics since the Lower Cretaceous has been characterized by a multi‐phase subduction and collision history with temporally and spatially‐variable, small‐scale plate configurations. A new shear‐wave velocity model of the Mediterranean upper mantle (MeRE2020), constrained by a very large set of over 200,000 broadband (8‐350 s), inter‐station, Rayleigh‐wave, phase‐velocity curves, illuminates the complex structure and fragmentation of the subducting slabs. Phase‐velocity maps computed using these measurements were inverted for depth‐dependent, shear‐wave velocities using a stochastic particle‐swarm‐optimization algorithm (PSO). The resulting three‐dimensional (3‐D) model makes possible an inventory of slab segments across the Mediterranean. Fourteen slab segments of 200‐800 km length along‐strike are identified. We distinguish three categories of subducted slabs: attached slabs reaching down to the bottom of the model; shallow slabs of shorter length in down‐dip direction, terminating shallower than 300 km depth; and detached slab segments. The location of slab segments are consistent with and validated by the intermediate‐depth seismicity, where it is present. The new high‐resolution tomography demonstrates the intricate relationships between slab fragmentation and the evolution of the relatively small and highly curved subduction zones and collisional orogens characteristic of the Mediterranean realm

Reakcija β-amino-α,γ-dicianokrotononitrila s acetofenonom: sinteza derivata piridina, piridazina i tiofena s antimikrobnim djelovanjem

Condensation of β-amino-α,γ-dicyanocrotononitrile (1) with acetophenone gave the 2-amino-4-phenylpenta-1,3-diene-1,1,3-tricarbonitrile (2). The latter product was used in a series of heterocyclization reactions when react with different reagents like diazonium salts, hydrazines, hydroxylamine and elemental sulfur to give pyridazine, pyrazole, isoxazole and thiophene derivatives, respectively. On the other hand, it gave pyridine derivatives with aromatic aldehydes followed by reaction with cyanomethylene reagents. The MIC values for the newly synthesized product were measured against E. coli, B. cereus, B. subtilis and C. albicansKondenzacijom β-amino-α,γ-dicijanokrotononitrila 1 s acetofenonom dobiven je 2-amino-4-fenilpenta-1,3-dien-1,1,3-trikarbonitril (2) koji je upotrebljen u reakcijama heterociklizacije s različitim reagensima poput diazonijevih soli, hidrazina, hidroksilamina i elementarnog sumpora pri čemu su nastali derivati piridazina, pirazola, izoksazola, odnosno tiofena. Spoj 2 je u reakciji s aromatskim aldehidima te naknadno sa cijanometilenima dao derivate piridina. Određene su MIC vrijednosti za novosintetizirane spojeve protiv E. coli, B. cereus, B. subtilis i C. albicans

Domestication syndrome is investigated by proteomic analysis between cultivated cassava (Manihot esculenta Crantz) and its wild relatives

Cassava (Manihot esculenta Crantz) wild relatives remain a largely untapped potential for genetic improvement. However, the domestication syndrome phenomena from wild species to cultivated cassava remain poorly understood. The analysis of leaf anatomy and photosynthetic activity showed significantly different between cassava cultivars SC205, SC8 and wild relative M. esculenta ssp. Flabellifolia (W14). The dry matter, starch and amylose contents in the storage roots of cassava cultivars were significantly more than that in wild species. In order to further reveal the differences in photosynthesis and starch accumulation of cultivars and wild species, the globally differential proteins between cassava SC205, SC8 and W14 were analyzed using 2-DE in combination with MALDI-TOF tandem mass spectrometry. A total of 175 and 304 proteins in leaves and storage roots were identified, respectively. Of these, 122 and 127 common proteins in leaves and storage roots were detected in SC205, SC8 and W14, respectively. There were 11, 2 and 2 unique proteins in leaves, as well as 58, 9 and 12 unique proteins in storage roots for W14, SC205 and SC8, respectively, indicating proteomic changes in leaves and storage roots between cultivated cassava and its wild relatives. These proteins and their differential regulation across plants of contrasting leaf morphology, leaf anatomy pattern and photosynthetic related parameters and starch content could contribute to the footprinting of cassava domestication syndrome. We conclude that these global protein data would be of great value to detect the key gene groups related to cassava selection in the domestication syndrome phenomena

HGF Mediates the Anti-inflammatory Effects of PRP on Injured Tendons

Platelet-rich plasma (PRP) containing hepatocyte growth factor (HGF) and other growth factors are widely used in orthopaedic/sports medicine to repair injured tendons. While PRP treatment is reported to decrease pain in patients with tendon injury, the mechanism of this effect is not clear. Tendon pain is often associated with tendon inflammation, and HGF is known to protect tissues from inflammatory damages. Therefore, we hypothesized that HGF in PRP causes the anti-inflammatory effects. To test this hypothesis, we performed in vitro experiments on rabbit tendon cells and in vivo experiments on a mouse Achilles tendon injury model. We found that addition of PRP or HGF decreased gene expression of COX-1, COX-2, and mPGES-1, induced by the treatment of tendon cells in vitro with IL-1β. Further, the treatment of tendon cell cultures with HGF antibodies reduced the suppressive effects of PRP or HGF on IL-1β-induced COX-1, COX-2, and mPGES-1 gene expressions. Treatment with PRP or HGF almost completely blocked the cellular production of PGE2 and the expression of COX proteins. Finally, injection of PRP or HGF into wounded mouse Achilles tendons in vivo decreased PGE2 production in the tendinous tissues. Injection of platelet-poor plasma (PPP) however, did not reduce PGE2 levels in the wounded tendons, but the injection of HGF antibody inhibited the effects of PRP and HGF. Further, injection of PRP or HGF also decreased COX-1 and COX-2 proteins. These results indicate that PRP exerts anti-inflammatory effects on injured tendons through HGF. This study provides basic scientific evidence to support the use of PRP to treat injured tendons because PRP can reduce inflammation and thereby reduce the associated pain caused by high levels of PGE2. © 2013 Zhang et al

Functional characterization of a melon alcohol acyl-transferase gene family involved in the biosynthesis of ester volatiles. Identification of the crucial role of a threonine residue for enzyme activity

Volatile esters, a major class of compounds contributing to the aroma of many fruit, are synthesized by alcohol acyl-transferases (AAT). We demonstrate here that, in Charentais melon (Cucumis melo var. cantalupensis), AAT are encoded by a gene family of at least four members with amino acid identity ranging from 84% (Cm-AAT1/Cm-AAT2) and 58% (Cm-AAT1/Cm-AAT3) to only 22% (Cm-AAT1/Cm-AAT4). All encoded proteins, except Cm-AAT2, were enzymatically active upon expression in yeast and show differential substrate preferences. Cm-AAT1 protein produces a wide range of short and long-chain acyl esters but has strong preference for the formation of E-2-hexenyl acetate and hexyl hexanoate. Cm-AAT3 also accepts a wide range of substrates but with very strong preference for producing benzyl acetate. Cm-AAT4 is almost exclusively devoted to the formation of acetates, with strong preference for cinnamoyl acetate. Site directed mutagenesis demonstrated that the failure of Cm-AAT2 to produce volatile esters is related to the presence of a 268-alanine residue instead of threonine as in all active AAT proteins. Mutating 268-A into 268-T of Cm-AAT2 restored enzyme activity, while mutating 268-T into 268-A abolished activity of Cm-AAT1. Activities of all three proteins measured with the prefered substrates sharply increase during fruit ripening. The expression of all Cm-AAT genes is up-regulated during ripening and inhibited in antisense ACC oxidase melons and in fruit treated with the ethylene antagonist 1-methylcyclopropene (1-MCP), indicating a positive regulation by ethylene. The data presented in this work suggest that the multiplicity of AAT genes accounts for the great diversity of esters formed in melon

The role of redox mechanisms in hepatic chronic wound healing and fibrogenesis

Under physiological conditions, intracellular and tissue levels of reactive oxygen species (ROS) are carefully controlled and employed as fine modulators of signal transduction, gene expression and cell functional responses (redox signaling). A significant derangement in redox homeostasis, resulting in sustained levels of oxidative stress and related mediators, plays a role in the pathogenesis of human diseases characterized by chronic inflammation, chronic activation of wound healing and tissue fibrogenesis, including chronic liver diseases. In this chapter major concepts and mechanisms in redox signaling will be briefly recalled to introduce a number of selected examples of redox-related mechanisms that can actively contribute to critical events in the natural history of a chronic liver diseases, including induction of cell death, perpetuation of chronic inflammatory responses and fibrogenesis. A major focus will be on redox-dependent mechanisms involved in the modulation of phenotypic responses of activated, myofibroblast-like, hepatic stellate cells (HSC/MFs), still considered as the most relevant pro-fibrogenic cells operating in chronic liver diseases

Transgenic Biofortification of the Starchy Staple Cassava (Manihot esculenta) Generates a Novel Sink for Protein

Although calorie dense, the starchy, tuberous roots of cassava provide the lowest sources of dietary protein within the major staple food crops (Manihot esculenta Crantz). (Montagnac JA, Davis CR, Tanumihardjo SA. (2009) Compr Rev Food Sci Food Saf 8:181–194). Cassava was genetically modified to express zeolin, a nutritionally balanced storage protein under control of the patatin promoter. Transgenic plants accumulated zeolin within de novo protein bodies localized within the root storage tissues, resulting in total protein levels of 12.5% dry weight within this tissue, a fourfold increase compared to non-transgenic controls. No significant differences were seen for morphological or agronomic characteristics of transgenic and wild type plants in the greenhouse and field trials, but relative to controls, levels of cyanogenic compounds were reduced by up to 55% in both leaf and root tissues of transgenic plants. Data described here represent a proof of concept towards the potential transformation of cassava from a starchy staple, devoid of storage protein, to one capable of supplying inexpensive, plant-based proteins for food, feed and industrial applications