Enkai Xu

Transcriptome/Degradome-Wide Discovery of MicroRNAs and Transcript Targets in Two Paulownia australis Genotypes

MicroRNAs (miRNAs) are involved in plant growth, development, and response to biotic and abiotic stresses. Most of the miRNAs that have been identified in model plants are well characterized, but till now, no reports have previously been published concerning miRNAs in Paulownia australis. In order to investigate miRNA-guided transcript target regulation in P. australis, small RNA libraries from two P. australis (diploids, PA2; and autotetraploids, PA4) genotypes were subjected to Solexa sequencing. As a result, 10,691,271 (PA2) and 10,712,733 (PA4) clean reads were obtained, and 45 conserved miRNAs belonging to 15 families, and 31 potential novel miRNAs candidates were identified. Compared with their expression levels in the PA2 plants, 26 miRNAs were up-regulated and 15 miRNAs were down-regulated in the PA4 plants. The relative expressions of 12 miRNAs were validated by quantitative real-time polymerase chain reaction. Using the degradome approach, 53 transcript targets were identified and annotated based on Gene Ontology and Kyoto Encyclopedia of Genes and Genomes pathway analysis. These targets were associated with development, stimulus response, metabolism, signaling transduction and biological regulation. Among them, 11 targets, including TCP transcription factors, auxin response factors, squamosa promoter-binding-like proteins, scarecrow-like proteins, L-type lectin-domain containing receptor kinases and zinc finger CCCH domain-containing protein, cleaved by four known miRNA family and two potentially novel miRNAs were found to be involved in regulating plant development, biotic and abiotic stresses. The findings will be helpful to facilitate studies on the functions of miRNAs and their transcript targets in Paulownia.

Genome-wide expression profiling of the transcriptomes of four Paulownia tomentosa accessions in response to drought

Paulownia tomentosa is an important foundation forest tree species in semiarid areas. The lack of genetic information hinders research into the mechanisms involved in its response to abiotic stresses. Here, short-read sequencing technology (Illumina) was used to de novo assemble the transcriptome on P. tomentosa. A total of 99,218 unigenes with a mean length of 949 nucleotides were assembled. 68,295 unigenes were selected and the functions of their products were predicted using Clusters of Orthologous Groups, Gene Ontology, and Kyoto Encyclopedia of Genes and Genomes annotations. Afterwards, hundreds of genes involved in drought response were identified. Twelve putative drought response genes were analyzed by quantitative real-time polymerase chain reaction. This study provides a dataset of genes and inherent biochemical pathways, which will help in understanding the mechanisms of the water-deficit response in P. tomentosa. To our knowledge, this is the first study to highlight the genetic makeup of P. tomentosa.

Transcriptome analysis of the variations between autotetraploid Paulownia tomentosa and its diploid using high-throughput sequencing

Timber properties of autotetraploid Paulownia tomentosa are heritable with whole genome duplication, but the molecular mechanisms for the predominant characteristics remain unclear. To illuminate the genetic basis, high-throughput sequencing technology was used to identify the related unigenes. 2677 unigenes were found to be significantly differentially expressed in autotetraploid P. tomentosa. In total, 30 photosynthesis-related, 21 transcription factor-related, and 22 lignin-related differentially expressed unigenes were detected, and the roles of the peroxidase in lignin biosynthesis, MYB DNA-binding proteins, and WRKY proteins associated with the regulation of relevant hormones are extensively discussed. The results provide transcriptome data that may bring a new perspective to explain the polyploidy mechanism in the long growth cycle of plants and offer some help to the future Paulownia breeding.

Transcriptome sequencing and comparative analysis of diploid and autotetraploid Paulownia australis

Paulownia is a fast-growing tree native to China. It can provide high-quality timber and a green environment for humans and has been planted in many countries. Compared with diploid Paulownia australis, autotetraploid P. australis exhibits predominance in terms of yields, qualities, growth, and resistance to stress. However, the molecular and biological mechanisms of its predominant characteristics are still unclear. We performed paired-end sequencing of the transcriptomes of diploid and autotetraploid P. australis plants using the Illumina/Solexa Genome Analyzer platform and compared changes in the expression of genes after chromosome doubling. We obtained a total of 48,445,468 and 46,437,758 high-quality reads from diploid P. australis (PA2) and autotetraploid P. australis (PA4), respectively. After assembly, 90,881 (PA2) and 88,566 (PA4) unigenes were obtained. The unigenes from both transcriptomes were then assembled into 93,300 all-unigenes, which included 50,892 clusters and 42,408 singletons. The all-unigene sequences were searched against public protein databases (Nr, Nt, Swiss-Prot, KOG, and KEGG) using BLASTX with a cutoff E-value of 10−5. Gene expression levels in the two libraries were compared, and a total of 8851 differentially expressed unigenes were identified. A large number of unigenes were found to be related to carbohydrate and energy metabolism, biosynthesis of cell wall, and stress tolerance. Many of the differentially expressed P. australis genes were attributable to chromosome doubling between the PA2 and PA4 plants. Functional analysis revealed variations in the mechanism of growth and stress resistance between PA2 and PA4.

Transcriptome-Wide Profiling and Expression Analysis of Diploid and Autotetraploid Paulownia tomentosa × Paulownia fortunei under Drought Stress

Paulownia is a fast-growing deciduous hardwood species native to China, which has high ecological and economic value. In an earlier study, we reported ploidy-dependent differences in Paulownia drought tolerance by the microscopic observations of the leaves. Autotetraploid Paulownia has a higher resistance to drought stress than their diploid relatives. In order to obtain genetic information on molecular mechanisms responses of Paulownia plants to drought, Illumina/Solexa Genome sequencing platform was used to de novo assemble the transcriptomes of leaves from diploid and autotetraploid Paulownia tomentosa × Paulownia fortunei seedlings (PTF2 and PTF4 respectively) grown under control conditions and under drought stress and obtained 98,671 nonredundant unigenes. A comparative transcriptome analysis revealed that hundreds of unigenes were predicted to be involved mainly in ROS-scavenging system, amino acid and carbohydrate metabolism, plant hormone biosynthesis and signal transduction, while these unigenes exhibited differential transcript alteration of the two accessions. This study provides a comprehensive map of how P. tomentosa × P. fortunei responds to drought stress at physiological and molecular levels, which may help in understanding the mechanisms involve in water-deficit response and will be useful for further study of drought tolerance in woody plants.

Identification of genes related to the phenotypic variations of a synthesized Paulownia (Paulownia tomentosa × Paulownia fortunei) autotetraploid

Paulownia is a fast-growing deciduous tree native to China. It has great economic importance for the pulp and paper industries, as well as ecological prominence in forest ecosystems. Paulownia is of much interest to plant breeder keen to explore new plant varieties by selecting on the basis of phenotype. A newly synthesized autotetraploid Paulownia exhibited advanced characteristics, such as greater yield, and higher resistance than the diploid tree. However, tissue-specific transcriptome and genomic data in public databases are not sufficient to understand the molecular mechanisms associated with genome duplication. To evaluate the effects of genome duplication on the phenotypic variations in Paulownia tomentosa×Paulownia fortunei, the transcriptomes of the autotetraploid and diploid Paulownia were compared. Using Illumina sequencing technology, a total of 82,934 All-unigenes with a mean length of 1109 bp were assembled. The data revealed numerous differences in gene expression between the two transcriptomes, including 718 up-regulated and 667 down-regulated differentially expressed genes between the two Paulownia trees. An analysis of the pathway and gene annotations revealed that genes involved in nucleotide sugar metabolism in plant cell walls were down-regulated, and genes involved in the light signal pathway and the biosynthesis of structural polymers were up-regulated in autotetraploid Paulownia. The differentially expressed genes may contribute to the observed phenotypic variations between diploid and autotetraploid Paulownia. These results provide a significant resource for understanding the variations in Paulownia polyploidization and will benefit future breeding work.

Transcriptome-wide profiling and expression analysis of two accessions of Paulownia australis under salt stress

Paulownia australis has important economic and ecological values. In this study, the morphological and physiological changes of the leaves in diploid and autotetraploid P. australis under salt stress were analyzed. To detect related genes and gain a comprehensive perspective on the molecular mechanisms underlying salt tolerance in P. australis, transcriptome-wide gene expression profiling was conducted in the leaves of the diploid and autotetraploid P. australis under control and salinity conditions, respectively. Evaluation of the responses against salinity stress revealed the superiority of autotetraploid over diploid in terms of salinity tolerance. Changes in physiological parameters in diploid P. australis (PA2) and tetraploid P. australis (PA4) plants in response to salt stress were measured. Transcriptome data revealed that many of the common unigenes which were involved in accumulation of compatible solutes, oxidative stress detoxification, ion homeostasis, and signal transduction showed significant differences between the two accessions in response to salt stress. A number of salt-responsive unigenes were identified in two accessions of P. australis under salt stress. Furthermore, the differentially expressed unigenes found to be common in both accessions may be useful genetic resources for further genetic improvement of Paulownia using transgenic approaches.

A comparison of the transcriptomes between diploid and autotetraploid Paulownia fortunei under salt stress

Paulownia is a tree species grown in many countries. Our previous study reveals that tetraploid Paulownia fortunei is more tolerant to salt stress than its corresponding diploid tree. To investigate the molecular mechanisms of salt stress tolerance in P. fortunei, the transcriptomes of normal and salt-stressed diploid and tetraploid were investigated. After assembling the clean reads, we obtained 130,842 unigenes. The unigenes were aligned against six public databases (Nr, Nt, Swiss-Prot, COG, KEGG, GO) to discover homologs and assign functional annotations. We retrieved 7983 and 15,503 differentially expressed unigenes (DEUs) between the normal and the salt-stressed diploid and tetraploid P. fortunei, respectively. We identified dozens of important DEUs including 3 related to photosynthesis, 10 related to plant growth and development and 11 related to osmolytes. Some of these DEUs were upregulated in tetraploid compared to diploid and others were upregulated under salt stress. Quantitative reverse transcriptase polymerase chain reaction verified the expression patterns of 15 unigenes. Our results provided insights into the molecular aspects why tetraploid is stronger and more energetic than diploid under saline environment. This study provides useful information for further studies on the molecular mechanisms of salt tolerance in other tree plants.