Non-heading Chinese cabbage (Brassica campestris L. ssp. chinensis Makino), an important vegetable crop in China, exhibits a typical sporophytic self-incompatibility (SI) system. To better understand the mechanism of SI response and identify potential candidate proteins involved in the SI system of this vegetable crop, the proteomic approach was taken to identify differential accumulating pistil proteins. Pistils were collected at 0 h and 2 h after self-pollination at anthesis in self-incompatible and compatible lines of non-heading Chinese cabbage, and total proteins were extracted and separated by two-dimensional gel electrophoresis (2-DE). A total of 25 protein spots that displayed differential abundance were identified by matrix-assisted laser desorption/ionisation-time of flight mass spectrometry (MALDI-TOF/TOF MS) and peptide mass fingerprinting (PMF). Among them, 22 protein spots were confidently established. The mRNA levels of the corresponding genes were detected by quantitative RT-PCR. The 22 identified protein spots are involved in energy metabolism (four), protein biosynthesis (three), photosynthesis (six), stress response and defence (five), and protein degradation (four). Among these potential candidate proteins, UDP-sugar pyrophosphorylase could be involved in sucrose degradation to influence pollen germination and growth. Glutathione S-transferases could be involved in pollen maturation, and affect pollen fertility. Senescence-associated cysteine protease, which is related to programmed cell death, could be mainly related to self pollen recognition of non-heading Chinese cabbage. The study will contribute to further investigations of molecular mechanism of sporophytic SI in Brassicaceae.
Viral disease, caused by turnip mosaic virus (TuMV), is considered the most destructive disease of non-heading Chinese cabbage (Brassica campestris ssp. chinensis Makino). Although several TuMV resistance loci/genes have been mapped or characterized in Brassica vegetables, the mechanism of molecular interaction between TuMV and non-heading Chinese cabbage is poorly understood. Additionally, TuMV response genes need to be identified. The objectives of this study were to identify differentially expressed genes during the incompatible interaction between TuMV and non-heading Chinese cabbage, and validate their expressions. A total of 200 transcript-derived fragments (TDFs) obtained by complementary DNAamplified fragment length polymorphism were recovered and sequenced. The results revealed that 176 (88.0%) TDFs produced specific sequences, among which 48 (27.3%) sequences were predicted with putative functions using NCBI BLAST. Among the 48 available TDFs, 22 (45.8%) sequences belonging to different functional groups were selected to monitor the changes in their expression in incompatible and compatible interactions by quantitative real-time polymerase chain reaction. To the best of our knowledge, this study provides the first global transcriptomic analysis of nonheading Chinese cabbage genes during an incompatible interaction. The results are expected to aid in characterizing TuMV response genes and clarifying the molecular mechanism of TuMV-host interaction.
Eukaryotic elongation factor Tu has been implicated in responses to heat stress and viral infection. In this study, the turnip mosaic virus (TuMV)-response gene BcLRK01, which encodes a leucine-rich repeat receptor-like kinase, was probed using the cDNA library of TuMV-infected leaves of non-heading Chinese cabbage (Brassica campestris ssp. chinensis). The BcEF-Tu gene, which encodes chloroplast elongation factor Tu, was obtained and verified by a yeast two-hybrid system to interact with the BcLRK01 gene. TuMV infection depressed the expression of this gene, whereas a heat stress induced its expression. Overexpression of BcEF-Tu enhanced the viability of Escherichia coli transformants under the heat stress. These results demonstrate that elongation factor BcEF-Tu responded to the TuMV infection and heat stress. This is the first report on chloroplast EF-Tu in non-heading Chinese cabbage which provides a theoretical basis for the functional research of EF-Tu.
The exocyst is a conserved protein complex, and required for vesicles tethering, fusion and polarized exocytosis. Exo70A1, the exocyst subunit, is essential for assembly of the exocyst complex. To better understand potential roles of Exo70A1 in non-heading Chinese cabbage (Brassica campestris ssp. chinensis), we obtained the full-length cDNA of Exo70A1 gene, which consisted of 1, 917 bp and encoded a protein of 638 amino acids. BlastX showed BcExo70A1 shared 94.9 % identity with Brassica oleracea var. acephala (AEI26267.1), and clustered into a same group with other homologues in Brassica oleracea var. acephala and Brassica napus. Subcellular localization analysis showed BcExo70A1 was localized to punctate structures in cytosol of onion epithelial cells.Results showed that BcExo70A1 was widely presented in stamens, young stems, petals, unpollinated pistils, roots and leaves of self compatible and incompatible plants. The transcripts of BcExo70A1 in non-heading Chinese cabbage declined during initial 1.5 h after incompatible pollination, while an opposite trend was presented after compatible pollination. Our study reveals that BcExo70A1 could play essential roles in plant growth and development, and is related to the rejection of self pollen in non-heading Chinese cabbage.
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