A Pectate Lyase Gene Plays a Critical Role in Xylem Vascular Development in Arabidopsis.

As a major component of the plant primary cell wall, structure changes in pectin may affect the formation of the secondary cell wall and lead to serious consequences on plant growth and development. Pectin-modifying enzymes including pectate lyase-like proteins (PLLs) participate in the remodeling of pectin during organogenesis, especially during fruit ripening. In this study, we used Arabidopsis as a model system to identify critical PLL genes that are of particular importance for vascular development. Four PLL genes, named AtPLL15, AtPLL16, AtPLL19, and AtPLL26, were identified for xylem-specific expression. A knock-out T-DNA mutant of AtPLL16 displayed an increased amount of pectin, soluble sugar, and acid-soluble lignin (ASL). Interestingly, the atpll16 mutant exhibited an irregular xylem phenotype, accompanied by disordered xylem ray cells and an absence of interfascicular phloem fibers. The xylem fiber cell walls in the atpll16 mutant were thicker than those of the wild type. On the contrary, AtPLL16 overexpression resulted in expansion of the phloem and a dramatic change in the xylem-to-phloem ratios. Altogether, our data suggest that AtPLL16 as a pectate lyase plays an important role during vascular development in Arabidopsis.

Expressing a Short Tandem Target Mimic (STTM) of miR164b/e-3p enhances poplar leaf serration by co-regulating the miR164-NAC module.

MicroRNAs (miRNAs) are short non-coding RNAs (21-24 nt) that play important roles in plant growth and development. The miR164 family is highly conserved in plants and the miR164-NAM/ATAF/CUC (NAC) module is validated to regulate leaf and flower development, lateral root initiation and stress response. However, our knowledge of its role in Populus remains limited. In this study, two mature miRNA species, miR164e-5p and miR164e-3p, were identified in Populus deltoides. Their nucleotide sequences were identical to those of miR164a/b/c/d/e-5p and miR164b/e-3p in P. tremula × P. alba clone 717-1B4 (hereinafter poplar 717), respectively. Transgenic plants of poplar 717, including overexpression lines (35S::pri-miR164e) and Short Tandem Target Mimic lines (STTM-miR164a-d,e-5p and STTM-miR164b/e-3p), were generated to study the roles of miR164e-5p and miR164e-3p in poplar. Compared with poplar 717, the leaf margins of 35S::pri-miR164e lines were smoother, the leaves of STTM-miR164b/e-3p line were more serrated, while the leaf morphology of STTM-miR164a-d,e-5p lines had no obvious change. In addition, both 35S::pri-miR164e and STTM-miR164b/e-3p plants had a dwarf phenotype. Expressions of miR164a-d,e-5p target genes, including PtaCUC2a, PtaCUC2b and PtaORE1, was significantly reduced in the apex of 35S::pri-miR164e lines. Green fluorescent protein (GFP) reporter assay showed that PtaCUC2a/2b and PtaORE1 were cleaved by miR164a-d,e-5p, and the cleavage was inhibited by STTM-miR164b/e-3p. Therefore, miR164b/e-3p may cooperate with miR164a-d,e-5p to regulate certain NAC members, such as PtaCUC2a/2b and PtaORE1, thereby regulating leaf development and plant growth in poplar. Our findings add new insights into the mechanisms by which the miR164-NAC module regulates plant development.

Poplar's Waterlogging Resistance Modeling and Evaluating: Exploring and Perfecting the Feasibility of Machine Learning Methods in Plant Science.

Floods, as one of the most common disasters in the natural environment, have caused huge losses to human life and property. Predicting the flood resistance of poplar can effectively help researchers select seedlings scientifically and resist floods precisely. Using machine learning algorithms, models of poplar's waterlogging tolerance were established and evaluated. First of all, the evaluation indexes of poplar's waterlogging tolerance were analyzed and determined. Then, significance testing, correlation analysis, and three feature selection algorithms (Hierarchical clustering, Lasso, and Stepwise regression) were used to screen photosynthesis, chlorophyll fluorescence, and environmental parameters. Based on this, four machine learning methods, BP neural network regression (BPR), extreme learning machine regression (ELMR), support vector regression (SVR), and random forest regression (RFR) were used to predict the flood resistance of poplar. The results show that random forest regression (RFR) and support vector regression (SVR) have high precision. On the test set, the coefficient of determination (R2) is 0.8351 and 0.6864, the root mean square error (RMSE) is 0.2016 and 0.2780, and the mean absolute error (MAE) is 0.1782 and 0.2031, respectively. Therefore, random forest regression (RFR) and support vector regression (SVR) can be given priority to predict poplar flood resistance.

Influence of Switchgrass TDIF-like Genes on Arabidopsis Vascular Development.

As a member of the CLAVATA3 (CLV3)/EMBRYO SURROUNDING REGION (CLE) family, the dodecapeptide tracheary element differentiation inhibitory factor (TDIF) has a major impact on vascular development in plants. However, the influence of polymorphisms in the TDIF peptide motif on activity remains poorly understood. The model plant, Arabidopsis provides a fast and effective tool for assaying the activity of TDIF homologs. Five TDIF homologs from a group of 93 CLE genes in switchgrass (Panicum virgatum), a perennial biomass crop, named PvTDIF-like (PvTDIFL) genes were studied. The expression levels of PvTDIFL1, PvTDIFL3 MR3, and PvTDIFL3 MR2 were relatively high and all of them were expressed at the highest levels in the rachis of switchgrass. The precursor proteins for PvTDIFL1, PvTDIFL3MR3, and PvTDIFL3MR2 contained one, three, and two TDIFL motifs, respectively. Treatments with exogenous PvTDIFL peptides increased the number of stele cells in the hypocotyls of Arabidopsis seedlings, with the exception of PvTDIFL_4p. Heterologous expression of PvTDIFL1 in Arabidopsis strongly inhibited plant growth, increased cell division in the vascular tissue of the hypocotyl, and disrupted the cellular organization of the hypocotyl. Although heterologous expression of PvTDIFL3 MR3 and PvTDIFL3 MR2 also affected plant growth and vascular development, PvTDIFL activity was not enhanced by the multiple TDIFL motifs encoded by PvTDIFL3 MR3 and PvTDIFL3 MR2. These data indicate that in general, PvTDIFLs are functionally similar to Arabidopsis TDIF but that the processing and activities of the PvTDIFL peptides are more complex.

Development of genic SSR marker resources from RNA-seq data in Camellia japonica and their application in the genus Camellia.

Camellia is a genus of flowering plants in the family Theaceae, and several species in this genus have economic importance. Although a great deal of molecular makers has been developed for molecular assisted breeding in genus Camellia in the past decade, the number of simple sequence repeats (SSRs) publicly available for plants in this genus is insufficient. In this study, a total of 28,854 potential SSRs were identified with a frequency of 4.63 kb. A total of 172 primer pairs were synthesized and preliminarily screened in 10 C. japonica accessions, and of these primer pairs, 111 were found to be polymorphic. Fifty-one polymorphic SSR markers were randomly selected to perform further analysis of the genetic relationships of 89 accessions across the genus Camellia. Cluster analysis revealed major clusters corresponding to those based on taxonomic classification and geographic origin. Furthermore, all the genotypes of C. japonica separated and consistently grouped well in the genetic structure analysis. The results of the present study provide high-quality SSR resources for molecular genetic breeding studies in camellia plants.

Construction of a high-density genetic map and its application for leaf shape QTL mapping in poplar.

MAIN CONCLUSION: High-quality and dense genetic maps were constructed, and leaf shape variation was dissected by QTL mapping in poplar. Species in the genus Populus, also known as poplars, are important woody species and considered model plants for perennial trees. High-density genetic maps are valuable genomic resources for population genetics. Here, we generated a high-quality and dense genetic map for an F1 poplar population using high-throughput NGS-based genotyping. A total of 92,097 high-quality SNP markers were developed by stringent filtering and identification. In total, 889 and 1650 SNPs formed the female and male genetic maps, respectively. To test the application of the genetic maps, QTL mapping of leaf shape was conducted for this F1 population. A total of nine parameters were scored for leaf shape variation in three different environments. Combining genetic maps and measurements of the nine leaf shape parameters, we mapped a total of 42 significant QTLs. The highest LOD score of all QTLs was 9.2, and that QTL explained the most (15.13%) trait variation. A total of nine QTLs could be detected in at least two environments, and they were located in two genomic regions. Within these two QTL regions, some candidate genes for regulating leaf shape were predicted through functional annotation. The successful mapping of leaf shape QTLs demonstrated the utility of our genetic maps. According to the performance of this study, we were able to provide high-quality and dense genetic maps and dissect the leaf shape variation in poplar.

Molecular and physiological responses in roots of two full-sib poplars uncover mechanisms that contribute to differences in partial submergence tolerance.

Poplar is a major afforestation tree species in flood-prone areas. Here, we compared molecular and physiological responses in the roots of two full-sib poplar clones, LS1 (flood-tolerant) and LS2 (flood-susceptive), subjected to stagnant flooding using transcript and metabolite profiling. LS1 displayed less phenotypic damage and superior leaf gas exchange and plant growth compared with those of LS2. We concluded that three characteristics might contribute to the differences in flood tolerance between LS1 and LS2. First, fermentation was initiated through lactic dehydrogenation in LS1 roots under flooding and subsequently dominated by alcohol fermentation. However, lactic dehydrogenase was persistently active in flooded LS2. Second, 13 differentially expressed genes associated with energy and O2 consumption processes under soil flooding had lower transcript levels in LS1 than those in LS2, which might contribute to better energy-/O2-saving abilities and behaviours in flood-tolerant LS1 than those in flood-susceptible LS2 under hypoxic stress. Third, LS1 possessed increased reactive oxygen species scavenging abilities compared with those of LS2 under edaphic flooding. Our data are a valuable contribution to understanding the mechanisms involved in the flood tolerance of poplar.

RNA-seq based transcriptomic analysis uncovers α-linolenic acid and jasmonic acid biosynthesis pathways respond to cold acclimation in Camellia japonica.

Camellia is a well-known ornamental flower native to Southeast of Asia, including regions such as Japan, Korea and South China. However, most species in the genus Camellia are cold sensitive. To elucidate the cold stress responses in camellia plants, we carried out deep transcriptome sequencing of 'Jiangxue', a cold-tolerant cultivar of Camellia japonica, and approximately 1,006 million clean reads were generated using Illumina sequencing technology. The assembly of the clean reads produced 367,620 transcripts, including 207,592 unigenes. Overall, 28,038 differentially expressed genes were identified during cold acclimation. Detailed elucidation of responses of transcription factors, protein kinases and plant hormone signalling-related genes described the interplay of signal that allowed the plant to fine-tune cold stress responses. On the basis of global gene regulation of unsaturated fatty acid biosynthesis- and jasmonic acid biosynthesis-related genes, unsaturated fatty acid biosynthesis and jasmonic acid biosynthesis pathways were deduced to be involved in the low temperature responses in C. japonica. These results were supported by the determination of the fatty acid composition and jasmonic acid content. Our results provide insights into the genetic and molecular basis of the responses to cold acclimation in camellia plants.

Effects of fragment traits, burial orientation and nutrient supply on survival and growth in Populus deltoides × P. simonii.

Clonal propagations of shoot or root fragments play pivotal roles in adaptation of clonal trees to environmental heterogeneity, i.e. soil nutrient heterogeneity and burials after disturbance. However, little is known about whether burial orientation and nutrient supply can alter the effects of fragment traits in Populus. Shoot and root fragments of Populus deltoides × P. simonii were subjected to burials in two different fragment diameters (0.5 and 2.0 cm), two fragment lengths (5 and 15 cm) and three burial orientations (horizontal, upward and downward). For the shoot fragments, survival and growth were significantly higher in the larger pieces (either in length or diameter) and the horizontal/upward burial position. On the contrary, the effect of burial position was reversed for the root fragments. Shoot/root fragments of 15 cm in length in horizontal burial position were then subjected to two different fragment diameters (0.5 and 2.0 cm) and four types of nutrient supplies (without nutrient, low frequency, high frequency and patchy). Growth of shoot fragments of 2.0 cm in diameter significantly increased in high frequency and patchy nutrient supplies than that of without nutrient treatment. These results suggest that burial orientation and nutrient supply could be employed in clonal propagations of cuttings, afforestation or regeneration in Populus.