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Pyruvate kinase (PK) is a key limited enzyme that regulates the glycolysis pathway, which play an important role in biological process including metabolism of basic substances in plants. Cyperus esculentus L. accumulating high levels of nutrients such as lipid/oil, starch, and sugar in tubers is an ideal model for investigating carbon metabolism and its regulatory mechanisms. A total of 7 CePK genes were identified by mining transcriptome data, including three plastidial CePKp (CePKpa, CePKpβ1 and CePKpβ2) and four cytosolic CePKc (CePKc1, CePKc2, CePKc3 and CePKc4). All CePK proteins encoded by CePK genes have the typical PK and PK_C domains. CePK proteins classified as the same type exhibit the similarity in sequence length, relative molecular weight, stability, and isoelectric point. CePKp proteins are hydrophilic proteins, and predicted to be located in the chloroplast. CePKc proteins are stable hydrophobic proteins, and predicted to be located in the cytoplasm. The tertiary structure prediction indicates that CePK proteins exist in the form of homologous tetramers. During the tuber germination and seedling establishing, the expression patterns of these CePK genes were significantly different, demonstrating that CePK genes may function differentially in regulating nutrient metabolism pathways in the tuber and seedling. The present data provide a new scientific basis for the in-depth analysis of the collaborative regulation mechanism underlying carbon flux and allocation and other biological processes in tuber germination and seedling establishing, as also benefiting breeding of high-yield and high-quality C. esculentus. [ABSTRACT FROM AUTHOR]

Perilla [Perilla frutescens (L.) Britt.] is a new type of edible-medicinal oil crop. The transcription factor WRI1 (WRINKLED1) is master regulator in transcriptional control of plant oil biosynthesis and accumulation. To explore the molecular mechanism of PfWRI1 transcription factor in regulating oil synthesis accumulation and stress response in perilla, PfWRI1 transcription factor was cloned from perilla by molecular cloning technique. The sequence characteristics and expression levels were analyzed using bioinformatics analysis tools and semi-quantitative reverse transcription polymerase chain reaction (sqRTPCR) and quantitative real-time reverse transcription polymerase chain reaction (qRT-PCR) techniques. The results showed that PfWRI1 transcription factor is a member of the AP2 (APETALA2) transcription factor family, which contains two AP2 con served domains and is localized at the nucleus. Random coil and α-helix were the main structural elements in the secondary structure of PfWRI1 protein. Phylogenetic analysis showed that the closest relatives to PfWRI1 are Sesamum indicum and Persea americana WRI1. The prediction of interacting proteins indicated that PfWRI1 protein might interact with LEC1 (LEAFYCOTYLEDON1), FATA (fatty acyl-acyl carrier protien thioesterase), FATB and PDAT (phospholipids: diacylglycerol acyltransferase). PfWRI1 expressed in all perilla organs/tissues tested, whereas their expression levels were different in these tissues. The expression level of PfWRI1 increased in the early developmental stage and then decreased, and which reached the peak l in seed at 30 days after flowering (DAF) with the seed maturity. It was suggested that PfWRI1 may play an important regulatory role in perilla seed oil synthesis. The expression of PfWRI1 transcription factor peaked after 12 hours under low temperature stress of perilla seedlings, indicating that PfWRI1 may be involved in the response to cold stress. This study lays a foundation for further research on the molecular mechanism and function of PfWRI1 in oil synthesis and accumulation and stress response of perilla, and provides a theoretical basis for genetic improvement and new variety breeding of perilla and other oil crops. [ABSTRACT FROM AUTHOR]

Perilla ( Perilla frutescens L.) is an important edible-medicinal oil crop, with its seed containing 46%-58% oil. Of perilla seed oil, α-linolenic acid (C18:3) accounts for more than 60%. Lysophosphatidic acid acyltransferase (LPAT) is one of the key enzymes responsible for triacylglycerol assembly in plant seeds, controlling the metabolic flow from lysophosphatidic acid to phosphatidic acid. In this study, the LPAT2 gene from the developing seeds of perilla was cloned and designated as PfLPAT2 . The expression profile of PfLPAT2 gene was examined in various tissues and different seed development stages of perilla (10, 20, 30, and 40 days after flowering, DAF) by quantitative real-time PCR (qRT-PCR). In order to detect the subcellular localization of PfLPAT2 protein, a fusion expression vector containing PfLPAT2 and GFP was constructed and transformed into Nicotiana benthamiana leaves by Agrobacterium -mediated infiltration. In order to explore the enzymatic activity and biological function of PfLPAT2 protein, an E . coli expression vector, a yeast expression vector and a constitutive plant overexpression vector were constructed and transformed into an E . coli mutant SM2-1, a wild-type Saccharomyces cerevisiae strain INVSc1, and a common tobacco ( Nicotiana tabacum , variety: Sumsun NN, SNN), respectively. The results showed that the PfLPAT2 open reading frame (ORF) sequence was 1 155 bp in length, encoding 384 amino acid residues. Functional structure domain prediction showed that PfLPAT2 protein has a typical conserved domain of lysophosphatidic acid acyltransferase. qRT-PCR analysis indicated that PfLPAT2 gene was expressed in all tissues tested, with the peak level in seed of 20 DAF of perilla. Subcellular localization prediction showed that PfLPAT2 protein is localized in cytoplasm. Functional complementation assay of PfLPAT2 in E . coli LPAAT mutant (SM2-1) showed that PfLPAT2 could restore the lipid biosynthesis of SM2-1 cell membrane and possess LPAT enzyme activity. The total oil content in the PfLPAT2 transgenic yeast was significantly increased, and the content of each fatty acid component changed compared with that of the non-transgenic control strain. Particularly, oleic acid (C18:1) in the transgenic yeast significantly increased, indicating that PfLPAT2 has a higher substrate preference for C18:1. Importantly, total fatty acid content in the transgenic tobacco leaves increased by about 0.42 times compared to that of the controls, with the C18:1 content doubled. The increased total oil content and the altered fatty acid composition in transgenic tobacco lines demonstrated that the heterologous expression of PfLPAT2 could promote host oil biosynthesis and the accumulation of health-promoting fatty acids (C18:1 and C18:3). This study will provide a theoretical basis and genetic elements for in-depth analysis of the molecular regulation mechanism of perilla oil, especially the synthesis of unsaturated fatty acids, which is beneficial to the genetic improvement of oil quality of oil crops.