Eight New Peptaibols from Sponge-Associated Trichoderma atroviride

Eight new and four known peptaibols were isolated from a strain of the fungus, Trichoderma atroviride (NF16), which was cultured from an Axinellid sponge collected from the East Mediterranean coast of Israel. The structures of the pure compounds were determined using HRMS, MS/MS and one- and two-dimensional NMR measurements. The isolated compounds belong to the trichorzianines, a family of 19-residue linear hydrophobic peptides containing a high proportion of α-aminoisobutyric acid (Aib), an acetylated N-terminus and a C-terminal amino alcohol. These new peptaibols exhibited antimicrobial activity against environmental bacteria isolated from the Mediterranean coast of Israel

Supplemental data for the publication: "The Role of Vitamin D in Emiliania huxleyi: A Microalgal Perspective on UV-B Exposure"

The following table contains raw transcriptomic data of Emiliania huxleyi CCMP3266 grown under continuous UV radiation or control conditions for 7, 10, and 13 days, obtained using the MARS-seq protocol (Keren-Shaul H. et al. 2013) Column A: E. huxleyi CCMP3266 gene locus IDs Column B: E. huxleyi CCMP3266 representative gene transcripts Columns C: Best matching transcripts in E. huxleyi CCMP1516 Columns D - U: Raw read counts Columns V - AM: DESew2 normalized read counts Columns AN - BE: rlog transformed read counts. Columns BF - FC: Each block of 6 columns represents the results of the differential gene expression analysis between two treatments (time point or UV/nonUV). Columns FD - FJ: Functional annotation

AUXIN RESPONSE FACTOR 2 Intersects Hormonal Signals in the Regulation of Tomato Fruit Ripening.

The involvement of ethylene in fruit ripening is well documented, though knowledge regarding the crosstalk between ethylene and other hormones in ripening is lacking. We discovered that AUXIN RESPONSE FACTOR 2A (ARF2A), a recognized auxin signaling component, functions in the control of ripening. ARF2A expression is ripening regulated and reduced in the rin, nor and nr ripening mutants. It is also responsive to exogenous application of ethylene, auxin and abscisic acid (ABA). Over-expressing ARF2A in tomato resulted in blotchy ripening in which certain fruit regions turn red and possess accelerated ripening. ARF2A over-expressing fruit displayed early ethylene emission and ethylene signaling inhibition delayed their ripening phenotype, suggesting ethylene dependency. Both green and red fruit regions showed the induction of ethylene signaling components and master regulators of ripening. Comprehensive hormone profiling revealed that altered ARF2A expression in fruit significantly modified abscisates, cytokinins and salicylic acid while gibberellic acid and auxin metabolites were unaffected. Silencing of ARF2A further validated these observations as reducing ARF2A expression let to retarded fruit ripening, parthenocarpy and a disturbed hormonal profile. Finally, we show that ARF2A both homodimerizes and interacts with the ABA STRESS RIPENING (ASR1) protein, suggesting that ASR1 might be linking ABA and ethylene-dependent ripening. These results revealed that ARF2A interconnects signals of ethylene and additional hormones to co-ordinate the capacity of fruit tissue to initiate the complex ripening process

Microarray analysis of <i>ARF2-OX</i> fruit.

<p>Gene expression was analyzed in WT at 42 dpa (mature green stage; WT-42G) and 53 dpa (red stage; WT-53R) and green (<i>ARF2-OX</i>-42G) and red (<i>ARF2-OX</i>-42R) patches from <i>ARF2-OX</i> fruit at 42 dpa by microarray analysis. Results are displayed as (A) a principal component analysis (PCA) and (B) a Venn diagram of the differentially expressed genes in the comparisons: <i>ARF2-OX</i>-42G to WT-42G; <i>ARF2-OX</i>-42R to WT-42G; and WT-53R to WT-42G. P-value<0.01 and FDR<0.05; dpa: days post anthesis.</p

Dimerization of the ARF2A protein and its interaction.

<p>ARF2A was cloned downstream of the DNA-binding domain (DB-ARF2A) and co-transformed into yeast with either (A) ARF2A cloned downstream of the activation domain (AD-ARF2A); or (B) ASR1 cloned downstream of the activation domain (AD-ASR1), yeast growth on media lacking leucine, tryptophan, histidine and adenine indicated positive protein-protein interactions. (C) Relative expression levels of <i>ASR1</i> in WT cv. MicroTom fruit at five developmental stages (IG: immature green; MG: mature green; Br: breaker; Or: orange; and R: red), error bars represent SE; statistical significance was evaluated using an ANOVA test (JMP software, SAS) with three biological repeats based on the average of three technical replicates, values indicated by the same letter (a,b,c) are not statistically significant, p-value<0.05. (D) A Bimolecular Fluorescence Complementation assay (BiFC) was carried out by transient expression in tobacco leaves; ARF2A was cloned downstream of the amino-terminal region of YFP (yellow fluorescent protein; YN-ARF2A) and ASR1 was cloned downstream of the carboxy-terminal region of YFP (YC-ASR1); leaf regions were examined for fluorescent signal by light and confocal fluorescence microscopy. Inset zoom region shows that the ARF2A-ASR1 interaction is nuclear localized. Scale bars in the light and confocal fluorescence microscopy represent 50 μm and 10 μm, respectively.</p

Metabolic analysis of <i>ARF2-OX</i> fruit.

<p>WT and <i>ARF2-OX</i> fruit were analyzed at 42 and 53 dpa by UPLC-qTOF-MS in positive mode, (A) results are visualized by a principle component analysis (PCA) plot; and displayed as histograms for (B) targeted flavonoids (upper row), and targeted glycoalkaloids (lower row). (C) Isoprenoids were analyzed in WT and <i>ARF2-OX</i> fruit at 42 and 53 dpa by HPLC. Grey bars represent WT, black bars <i>ARF2-OX</i> green/yellow patches and white bars <i>ARF2-OX</i> red patches. Error bars represent SE. Statistical significance was evaluated using a student’s t-test, *p-value<0.05; dpa: days post anthesis.</p

Phenotype and expression levels of <i>ARF2</i> genes in <i>ARF2as</i> transgenic lines.

<p>Fruit of <i>ARF2as</i> lines were analysed for relative expression levels by qRT-PCR of (A) <i>ARF2A;</i> and (B) <i>ARF2B</i>. (C-D) The ripening of <i>ARF2as</i> fruit is delayed as compared to WT. (D-E) <i>ARF2as</i> fruit were parthenocarpic or nearly parthenocarpic with reduced number of seeds. (F) Principle component analysis (PCA) plot from untargeted analysis of metabolites. DPA: days post anthesis; error bars represent SE; Statistical significance was evaluated using a student’s t-test, **p-value<0.01.</p

<i>ARF2A</i> expression in tomato fruit.

<p>Relative expression levels of <i>ARF2A</i> analyzed by qRT-PCR, in (A) WT fruit at five developmental stages; (B) <i>rin</i> and <i>nor</i> mutants; (C) <i>TAGL1</i> over-expressing fruit (<i>35S</i>:<i>TAGL1</i>); and (D) <i>nr</i> mutant. Relative expression levels of <i>ARF2A</i> in fruit at three developmental stages, treated with (E) 1-MCP; (F) ethylene; (H) NAA; and (J) ABA. Relative expression levels of <i>ARF2A</i> in fruit at the MG stage, at 0, 2, 4 and 6 days post-treatment with (G) ethrel; (I) NAA; and (K) ABA. Error bars represent SE. Statistical significance was evaluated using a student’s t-test, *p-value<0.05, **p-value<0.01 and ***p-value<0.001; dpa: days post anthesis; IG: immature green; MG: mature green; Br: breaker; Or: orange; and R: red.</p

Altering ethylene signaling in <i>ARF2-OX</i> transgenic fruit.

<p><i>ARF2-OX</i> fruit at the mature green (MG) stage, before the visual appearance of patches, were treated with either (A) ethrel, or (B) 1-MCP, and phenotypes were observed at (A) 10 and 16, or (B) 7 and 10 DPT. (C) Ethylene emission was measured from WT and <i>ARF2-OX</i> fruit harvested at the MG stage, every 1–3 days for 16 days, the red bars and arrows indicate the breaker stage. Error bars represent SE. DPT: days post treatment.</p