Genome-Wide Analysis of Transcription Factor R2R3-MYB Gene Family and Gene Expression Profiles during Anthocyanin Synthesis in Common Walnut (Juglans regia L.).

The R2R3-MYB gene family, encoding plant transcriptional regulators, participates in many metabolic pathways of plant physiology and development, including flavonoid metabolism and anthocyanin synthesis. This study proceeded as follows: the JrR2R3-MYB gene family was analyzed genome-wide, and the family members were identified and characterized using the high-quality walnut reference genome "Chandler 2.0". All 204 JrR2R3-MYBs were established and categorized into 30 subgroups via phylogenetic analysis. JrR2R3-MYBs were unevenly distributed over 16 chromosomes. Most JrR2R3-MYBs had similar structures and conservative motifs. The cis-acting elements exhibit multiple functions of JrR2R3-MYBs such as light response, metabolite response, and stress response. We found that the expansion of JrR2R3-MYBs was mainly caused by WGD or segmental duplication events. Ka/Ks analysis indicated that these genes were in a state of negative purifying selection. Transcriptome results suggested that JrR2R3-MYBs were widely entangled in the process of walnut organ development and differentially expressed in different colored varieties of walnuts. Subsequently, we identified 17 differentially expressed JrR2R3-MYBs, 9 of which may regulate anthocyanin biosynthesis based on the results of a phylogenetic analysis. These genes were present in greater expression levels in 'Zijing' leaves than in 'Lvling' leaves, as revealed by the results of qRT-PCR experiments. These results contributed to the elucidation of the functions of JrR2R3-MYBs in walnut coloration. Collectively, this work provides a foundation for exploring the functional characteristics of the JrR2R3-MYBs in walnuts and improving the nutritional value and appearance quality of walnuts.

Genome-wide identification analysis of the 4-Coumarate: CoA ligase (4CL) gene family expression profiles in Juglans regia and its wild relatives J. Mandshurica resistance and salt stress.

Persian walnut (Juglans regia) and Manchurian walnut (Juglans mandshurica) belong to Juglandaceae, which are vulnerable, temperate deciduous perennial trees with high economical, ecological, and industrial values. 4-Coumarate: CoA ligase (4CL) plays an essential function in plant development, growth, and stress. Walnut production is challenged by diverse stresses, such as salinity, drought, and diseases. However, the characteristics and expression levels of 4CL gene family in Juglans species resistance and under salt stress are unknown. Here, we identified 36 Jr4CL genes and 31 Jm4CL genes, respectively. Based on phylogenetic relationship analysis, all 4CL genes were divided into three branches. WGD was the major duplication mode for 4CLs in two Juglans species. The phylogenic and collinearity analyses showed that the 4CLs were relatively conserved during evolution, but the gene structures varied widely. 4CLs promoter region contained multiply cis-acting elements related to phytohormones and stress responses. We found that Jr4CLs may be participated in the regulation of resistance to anthracnose. The expression level and some physiological of 4CLs were changed significantly after salt treatment. According to qRT-PCR results, positive regulation was found to be the main mode of regulation of 4CL genes after salt stress. Overall, J. mandshurica outperformed J. regia. Therefore, J. mandshurica can be used as a walnut rootstock to improve salt tolerance. Our results provide new understanding the potential functions of 4CL genes in stress tolerance, offer the theoretical genetic basis of walnut varieties adapted to salt stress, and provide an important reference for breeding cultivated walnuts for stress tolerance.

Genome-wide identification, characterization, and expression pattern of the late embryogenesis abundant (LEA) gene family in Juglans regia and its wild relatives J. mandshurica.

BACKGROUND: Late Embryogenesis Abundant (LEA) proteins are a class of proteins associated with plant stress resistance. Two Juglans species, Juglans regia and J. mandshurica, are both diploid (2n = 32), monoecious perennial economic tree species with high edible, pharmaceutical, and timber value. The identification, characterization, and expression patterns of LEA proteins in J. regia and its wild relative, J. mandshurica, would not only provide the genetic basis of this gene family, but it would also supply clues for further studies of the evolution and regulating mechanisms of LEA proteins in other tree species. RESULTS: In this study, we identified 25 and 20 members of the LEA gene family in Juglans regia and its wild relative, Juglans mandshurica, respectively. The results of phylogenetic analysis showed that the LEA members were divided into eight main subgroups. Predictions of their physicochemical properties showed the variable characteristics of LEA proteins, and the subcellular localization analysis indicated that most LEA proteins are localized in the nucleus. Chromosomal localization analysis and gene replication pattern prediction indicated that WGD is the predominant duplication mode of LEA genes. The results of the comparative analysis indicated a high level of collinearity between the two Juglans species. Analysis of cis-acting elements indicated that LEA genes had a relatively wide range of responses to abiotic stresses and phytohormonal processes, particularly in two phytohormones, methyl jasmonate and abscisic acid. Transcriptome profiling and qRT-PCR experiments showed that JrLEAs are commonly expressed in leaves, green husks, and male and female flowers, and most JmLEAs are more highly expressed in male flowers. We also hypothesized that JrLEAs are involved in the process of anthracnose resistance. Anthracnose-resistant varieties of JrLEAs presented relatively high expression levels at later stages. CONCLUSION: In this study, we provide a theoretical basis for the functional study of LEA genes in J. regia and J. mandshurica. Analysis of cis-acting elements and gene expression indicated that JrLEAs and JmLEAs play important roles in resistance to biotic stresses in these species.