Functional characterization of alpha-synuclein protein with antimicrobial activity.

Alpha-synuclein (α-Syn), a small (14 kDa) protein associated with Parkinson's disease, is abundant in human neural tissues. α-Syn plays an important role in maintaining a supply of synaptic vesicles in presynaptic terminals; however, the mechanism by which it performs this function are not well understood. In addition, there is a correlation between α-Syn over-expression and upregulation of an innate immune response. Given the growing body of literature surrounding antimicrobial peptides (AMPs) in the brain, and the similarities between α-Syn and a previously characterized AMP, Amyloid-ß, we set out to investigate if α-Syn shares AMP-like properties. Here we demonstrate that α-Syn exhibits antibacterial activity against Escherichia coli and Staphylococcus aureus. In addition, we demonstrate a role for α-Syn in inhibiting various pathogenic fungal strains such as Aspergillus flavus, Aspergillus fumigatus and Rhizoctonia solani. We also analyzed localizations of recombinant α-Syn protein in E. coli and Candida albicans. These results suggest that in addition to α-Syn's role in neurotransmitter release, it appears to be a natural AMP.

Isolation of three B-box zinc finger proteins that interact with STF1 and COP1 defines a HY5/COP1 interaction network involved in light control of development in soybean.

LONG HYPOCOTYL5 (HY5) and STF1 (Soybean TGACG-motif binding Factor 1) are two related bZIP transcription factors that play a positive role in photomorphogenesis and hormonal signaling. In this study, we compared full length STF1 and truncated STF1 overexpression lines and found that the C-terminal 133 amino acids (194-306) possess all the HY5-like function in Arabidopsis. The STF1-DC1 mutant (1-306), with a 20 amino acid deletion at the carboxy terminus, failed to complement the hy5 mutant phenotype, which suggests an intact C-terminus is required for STF1 function. To understand the role of the C-terminal domain in photomorphogenesis we used a yeast two-hybrid screen to isolate proteins that bind to the STF1 C-terminus. We isolated three soybean cDNAs encoding the zinc-finger proteins GmSTO, GmSTH, and GmSTH2, which interact with STF1. These proteins belong to a family of B-box zinc finger proteins that include Arabidopsis SALT TOLERANCE (STO) and STO HOMOLOG (STH) and STH2, which play a role in light-dependent development and gene expression. The C-terminal 63 amino acids of STF1, containing a leucine zipper and the two N-terminal B-boxes, contains the domain involved in interactions between STF1 and GmSTO. In addition, we identified an interaction between soybean COP1 (GmCOP1) and GmSTO and GmSTH, as well as STF1, which strongly suggests the presence of a similar regulatory circuit for light signaling in soybean as in Arabidopsis. This study shows that photomorphogenic control requires complex molecular interactions among several different classes of transcription factors such as bZIP, B-box factors, and COP1, a ubiquitin ligase.

Overexpression of Arabidopsis NADPH-dependent thioredoxin reductase C (AtNTRC) confers freezing and cold shock tolerance to plants.

Overexpression of AtNTRC (AtNTRC(OE)) in Arabidopsis thaliana led to a freezing and cold stress tolerance, whereas a knockout mutant (atntrc) showed a stress-sensitive phenotype. Biochemical analyses showed that the recombinant AtNTRC proteins exhibited a cryoprotective activity for malate dehydrogenase and lactic dehydrogenase. Furthermore, conclusive evidence of its interaction with nucleic acids in vitro is provided here on the basis of gel shift and electron microscopy analysis. Recombinant AtNTRC efficiently protected RNA and DNA from RNase A and metal catalyzed oxidation damage, respectively. The C-terminal thioredoxin domain is required for the nucleic acid-protein complex formation. From these results, it can be hypothesized that AtNTRC, which is known to be an electron donor of peroxiredoxin, contributes the stability of macromolecules under cold stress.

Ethylene: A Modulator of the Phytohormone-Mediated Insect Herbivory Network in Plants.

Plants have evolved to establish insect herbivory defences by modulating their metabolism, growth, and development. Precise networks of phytohormones are essential to induce those herbivory defences. Gaseous phytohormone ET plays an important role in forming herbivory defences. Its role in insect herbivory is not fully understood, but previous studies have shown that it can both positively and negatively regulate herbivory. This review presents recent findings on crosstalk between ET and other phytohormones in herbivory responses. Additionally, the use of exogenous ETH treatment to induce ET in response to herbivory is discussed.

Comparative Analysis of Primary and Secondary Metabolites in Different In Vitro Tissues of Narcissus tazetta var. chinensis.

Narcissus tazetta var. chinensis is a perennial monocot plant that is well known for its pharmaceutical and ornamental uses. This study aimed to understand the changes in the primary and secondary metabolites in different in vitro tissues of N. tazetta (callus, adventitious root, and shoot) using high-performance liquid chromatography and gas chromatography time-of-flight mass spectrometry. In addition, to optimize the most efficient in vitro culture methods for primary and secondary metabolite production, N. tazetta bulbs were used as explants and cultivated in Murashige and Skoog (MS) medium containing different hormones at various concentrations. In addition, the present study found suitable hormonal concentrations for callus, adventitious root, and shoot induction and analyzed the primary and secondary metabolites. The MS medium supplemented with 1.0 mg L-1 dicamba, 3.0 mg L-1 indole-3-butyric acid (IBA), and 3.0 mg L-1 6-benzylaminopurine (BAP) was the most efficient media for callus, adventitious root, and shoot induction in N. tazetta. The tissue induced in this medium was subjected to primary (amines, amino acids, organic acids, sugars, and sugar alcohols) and secondary metabolite (galantamine and phenolic acids) analysis. The shoots and roots showed the highest amounts of metabolites. This study showed that bulb in vitro culture can be an efficient micropropagation method for N. tazetta and the production of primary and secondary metabolites, offering implications for the mass production of primary and secondary metabolite compounds from N. tazetta tissues generated in vitro.

Isolation of CONSTANS as a TGA4/OBF4 interacting protein.

Members of the TGA family of basic domain/leucine zipper transcription factors regulate defense genes through physical interaction with NON-EXPRESSOR OF PR1 (NPR1). Of the seven TGA family members, TGA4/octopine synthase (ocs)-element-binding factor 4 (OBF4) is the least understood. Here we present evidence for a novel function of OBF4 as a regulator of flowering. We identified CONSTANS (CO), a positive regulator of floral induction, as an OBF4-interacting protein, in a yeast two-hybrid library screen. OBF4 interacts with the B-box region of CO. The abundance of OBF4 mRNA cycles with a 24 h rhythm under both long-day (LD) and short-day (SD) conditions, with significantly higher levels during the night than during the day. Electrophoretic mobility shift assays revealed that OBF4 binds to the promoter of the FLOWERING LOCUS T (FT) gene, a direct target of CO. We also found that, like CO and FT, an OBF4:GUS construct was prominently expressed in the vascular tissues of leaf, indicating that OBF4 can regulate FT expression through the formation of a protein complex with CO. Taken together, our results suggest that OBF4 may act as a link between defense responses and flowering.

DNA-binding study identifies C-box and hybrid C/G-box or C/A-box motifs as high-affinity binding sites for STF1 and LONG HYPOCOTYL5 proteins.

LONG HYPOCOTYL5 (HY5) is a bZIP (basic leucine zipper) transcription factor that activates photomorphogenesis and root development in Arabidopsis (Arabidopsis thaliana). Previously, STF1 (soybean [Glycine max] TGACG-motif binding factor 1), a homologous legume protein with a RING-finger motif and a bZIP domain, was reported in soybean. To investigate the role of STF1, the phenotypes of transgenic Arabidopsis plants overexpressing STF1 and HY5 were compared. In addition, the DNA-binding properties of STF1 and HY5 were extensively studied using random binding site selection and electrophoretic mobility shift assay. Overexpression of STF1 in the hy5 mutant of Arabidopsis restored wild-type photomorphogenic and root development phenotypes of short hypocotyl, accumulation of chlorophyll, and root gravitropism with partial restoration of anthocyanin accumulation. This supports that STF1 is a homolog of HY5 with a role in light and hormone signaling. The DNA-binding properties of STF1 and HY5 are shown to be similar to each other in recognizing many ACGT-containing elements with a consensus sequence motif of 5'-(G/A)(G/A) TGACGT(C/G/A)(A/T/G)-3'. The motif represents a characteristically strong preference for flanking sequence to TGACGT and a larger sequence than the sequences recognized by the G-box binding factor and TGA protein families. The finding of C-box, hybrid C/G-, and C/A-boxes as high-affinity binding sites over the G-box and parameters associated with HY5 recognition define the criteria of HY5/STF1 protein-DNA interaction in the promoter regions. This study helps to predict the precise in vivo binding sites of the HY5 protein from the vast number of putative HY5 genomic binding sites analyzed by chromatin immunoprecipitation on chip.