Novel insight into anthocyanin metabolism and molecular characterization of its key regulators in Camellia sasanqua.

Flower color is a trait that affects the ornamental value of a plant. Camellia sasanqua is a horticultural plant with rich flower color, but little is known about the regulatory mechanism of color diversity in this plant. Here, the anthocyanin profile of 20 C. sasanqua cultivars revealed and quantified 11 anthocyanin derivatives (five delphinidin-based and six cyanidin-based anthocyanins) for the first time. Cyanidin-3-O-(6-O-(E)-p-coumaroyl)-glucoside was the main contributor to flower base color, and the accumulation of cyanidin and delphinidin derivatives differed in the petals. To further explore the molecular mechanism of color divergence, a transcriptome analysis was performed using C. sasanqua cultivars 'YingYueYe', 'WanXia', 'XueYueHua', and'XiaoMeiGui'. The co-expression network related to differences in delphinidin and cyanidin derivatives accumulation was identified. Eleven candidate genes encoding key enzymes (e.g., F3H, F3'H, and ANS) were involved in anthocyanin biosynthesis. Moreover, 27 transcription factors were screened as regulators of the two types of accumulating anthocyanins. The association was suggested by correlation analysis between the expression levels of the candidate genes and the different camellia cultivars. We concluded that cyanidin and delphinidin derivatives are the major drivers of color diversity in C. sasanqua. This finding provides valuable resources for the study of flower color in C. sasanqua and lays a foundation for genetic modification of anthocyanin biosynthesis.

Floral organ transcriptome in Camellia sasanqua provided insight into stamen petaloid.

BACKGROUND: The cultivated Camellia sasanqua forms a divergent double flower pattern, and the stamen petaloid is a vital factor in the phenomenon. However, the regulation mechanism remains largely unclear. RESULTS: Here, a comprehensive comparative transcriptome analysis of the wild-type, "semi-double", "peony double", and "rose double" was performed. The cluster analysis of global gene expression level showed petal and stamen difficulty separable in double flower. The crucial pathway and genes related to double flower patterns regulation were identified by pairwise comparisons and weighted gene coexpression network (WGCNA). Divergent genes expression, such as AUX1 and AHP, are involved in plant hormone signaling and photosynthesis, and secondary metabolites play an important role. Notably, the diversity of a petal-specific model exhibits a similar molecular signature to the stamen, containing extensin protein and PSBO1, supporting the stamen petaloid point. Moreover, the expansion of class A gene activity influenced the double flower formation, showing that the key function of gene expression was probably demolished. CONCLUSIONS: Overall, this work confirmed the ABCE model and provided new insights for elucidating the molecular signature of double formation.

Effect of Residual Stress Induced by Different Cooling Methods in Heat Treatment on the Fatigue Crack Propagation Behaviour of GH4169 Disc.

In this study, the effects of residual stress induced by three different cooling methods during heat treatment on the crack propagation behaviour of the GH4169 disc were investigated. Different levels of stress fields were induced to the specially designed discs by using air cooling (AC), air jetting cooling (AJC) and water quenching (WQ) methods and were quantitated by numerical simulation. These discs were then subjected to prefabricated cracking, and crack propagation tests were conducted on a spin tester with two load spectrums. Crack growth behaviour was depicted via the surface replica technique and fracture morphology. Regarding the linear superposition of residual stress and centrifugal force, the crack propagation behaviour of different discs was simulated using the FRANC3D software. AJC and WQ introduced compressive residual stress (-259 MPa and -109 MPa, respectively) into the disc compared with the AC method (about -1.5 MPa). The AJC method increases the crack propagation life of the disc by introducing residual compressive stress into the area near the surface of the central hole to inhibit the opening of the crack surface. When the fatigue load was low, this inhibition effect was more significant.

Multi-Approach Analysis Reveals Pathways of Cold Tolerance Divergence in Camellia japonica.

Understanding the molecular mechanism of the cold response is critical to improve horticultural plant cold tolerance. Here, we documented the physiological, transcriptome, proteome, and hormonal dynamics to cold stress in temperate genotype (Tg) and subtropical genotype (Sg) populations of Camellia japonica. Tg C. japonica suffered minimal osmotic and oxidative damage compared to Sg C. japonica under the same cold treatment. Transcriptional and translational differences increased under the cold treatment, indicating that Tg C. japonica was affected by the environment and displayed both conserved and divergent mechanisms. About 60% of the genes responding to cold had similar dynamics in the two populations, but 1,896 transcripts and 455 proteins differentially accumulated in response to the cold between Tg and Sg C. japonica. Co-expression analysis showed that the ribosomal protein and genes related to photosynthesis were upregulated in Tg C. japonica, and tryptophan, phenylpropanoid, and flavonoid metabolism were regulated differently between the two populations under cold stress. The divergence of these genes reflected a difference in cold responsiveness. In addition, the decrease in the abscisic acid (ABA)/gibberellic acid (GA) ratio regulated by biosynthetic signal transduction pathway enhanced cold resistance in Tg C. japonica, suggesting that hormones may regulate the difference in cold responsiveness. These results provide a new understanding of the molecular mechanism of cold stress and will improve cold tolerance in horticultural plants.

Overexpression of LtKNOX1 from Lilium tsingtauense in Nicotiana benthamiana affects the development of leaf morphology.

Leaves are the main vegetative organs of the aboveground part of plants and play an important role in plant morphogenesis. KNOTTED-LIKE HOMEOBOX (KNOX) plays a crucial role in regulating leaf cell fate and maintaining leaf development. In this study, we analyzed LtKNOX1 from Lilium tsingtauense and illustrated its function in transgenic plants. Tissue-specific expression analysis indicated that LtKNOX1 was highly expressed in stems, young flower buds, and shoot apical meristems (SAMs). Ectopic overexpression of LtKNOX1 in Nicotiana benthamiana suggested that transformants with mild phenotypes were characterized by foliar wrinkles and mildly curled leaves; transformants with intermediate phenotypes showed severely crimped blades and narrow leaf angles, and the most severe phenotypes lacked normal SAMs and leaves. Moreover, the expression levels of genes involved in the regulation of KNOX in transgenic plants were detected, including ASYMMETRIC LEAVES1, PIN-FORMED 1, GA20-oxidase, CUP-SHAPED COTYLEDON 2, CLAVATA 1 and WUSCHEL(WUS), and the expression of other genes were down-regulated except WUS. This study contributes to our understanding of the LtKNOX1 function.

Temporal transcriptome profiling reveals candidate genes involved in cold acclimation of Camellia japonica (Naidong).

Cold is a common problem that limits the distribution of Camellia. Camellia japonica (Naidong) is the northernmost species of camellia in China, which is a Tertiary remnant species that can adapt to large changes in temperature. An analysis of the transcriptional response of C. japonica (Naidong) to cold is very important for the planting and distribution of camellia. In this study, the rate of H2O2 levels, electrolyte leakage, chlorophyll and sugar content had a higher degree of cold response during 12-72 h period, than other periods (0-12h, 72h-120h) in C. japonica (Naidong) response to cold treatment. We constructed the first full-length C. japonica (Naidong) transcriptome and identified 4544 significantly differentially expressed genes (DEGs). A weighted gene coexpression network analysis showed that carbon metabolism, lipid metabolism, and transcription factors played important roles in the resistance of C. japonica (Naidong) to cold stress, and three hub transcription factor regulatory networks were constructed. In addition, overexpressing CjRAV1 led to cold sensitivity in Arabidopsis thaliana, thus CjRAV1 likely plays a negative regulatory role during cold stress in Camellia japonica. This study deepens our understanding of the regulatory mechanism of C. japonica (Naidong) under cold stress and will benefit genetic improvement of camellia.

CBF transcription factors involved in the cold response of Camellia japonica (Naidong).

CBFs belong to the ERF subfamily of the AP2 supergene family and often play an important role in the cold acclimation of temperate plants. However, the role of CBFs in Camellia japonica (Naidong), the only Camellia japonica population found in the temperate zones of China, remains unclear. It is very important to study the genetic composition of C. japonica (Naidong) to adapt to low temperature for Camellia species. Using full-length transcriptome data, we identified four CjCBF genes that respond to cold stress and analyzed their evolutionary relationships, domains, and expression patterns. The phylogeny of CBFs of 19 angiosperms divided the genes into three categories, and the four CjCBFs belong to a small subcluster. The strong response of CjCBF1 to cold treatment and its sustained high level of expression indicated that it plays an important role in the process of cold acclimation. A yeast two-hybrid assay revealed an interaction between CjCBF1, CjCBF2, and CjCBF5, and subcellular localization confirmed this finding. The expression of CjCBFs was tissue-specific: CBF1 was mainly expressed in leaves, and CBF3 was mainly expressed in stem. The responses of the four CjCBFs to drought and high temperature and the effect of light were also characterized. Our study provides new insight into the role of CBFs in the cold response in C. japonica (Naidong).

The complete chloroplast genome of the subtropical species Camellia japonica 'Huaheling'.

Camellia japonica 'Huaheling' is a rare subtropical Camellia species in China with high ornamental and medicinal value. The complete chloroplast genome of C. japonica 'Huaheling' is a 157,001-bp circular DNA molecule containing a large single-copy region (LSC, 86,704 bp), a small single-copy region (SSC, 18,393 bp), and two inverted repeat sequences (IR). Of the 131 genes identified, 86 are protein-coding genes, 8 are rRNA genes, and 37 are tRNA genes. A total of 54 simple sequence repeats (SSRs) were identified in the chloroplast genome. The phylogenetic analysis showed that C. japonica 'Huaheling' is clustered with C. japonica. This work provides valuable information for future study of the evolution and genetic diversity of C. japonica 'Huaheling.'