Transcriptional Regulation And Dissection Of Allelic Variations Of Gibberellin Response In Populus Tomentosa Transcriptional Regulation And Dissection Of Allelic Variations OfGibberellin Response In Populus Tomentosa

Forest is a very important part of terrestrial ecosystem, with a wide range of industrial uses and important ecological value. Recent years, the demand for timber is increasing, and the requirement for the quality of wood is higher and higher. Therefore, one of the main objectives of forest breeding is to increase the yield and improve the quality of wood. Gibberellins are important plant hormones, which play important roles in the growth and development of plants. In trees, GAs can affect tree growth, photosynthesis and wood formation, but specific mechanisms of these effects need to be further explored. Therefore, it is necessary to carry out the research on the transcriptional regulation of the response to GAs in forest trees. Thus, this study used the model tree of woody plants as the material to systematically study of the transcriptional regulation of GA response by using RNA-seq. This study also dissected the allelic variations within GA-responsive genes and noncoding RNA genes by using association mapping and identified SNPs significantly associated with photosynthetic, growth and wood quality traits, revealing the genetic regulatory network of lncRNA-miRNA-mRNA on tree growth and wood quality traits. This study provides a theoretical basis and technical support for the research of genetic regulation of GA response and molecular marker assisted breeding in forest trees. The main results and conclusions in this study as follows:1. The physiological and biochemical indexes, photosynthesis, growth and wood quality characteristics of Populus tomentosa were determined after GA treatment. The results showed that the total protein content, sucrose phosphate synthase （SPS） activity, peroxidase （POD） activity and malondialdehyde （MDA） content were significantly affected within 24 h after GA treatment. The physiological changes were the most obvious at 6 h after GA treatment. Four weeks after treatment, GA can significantly improve the Pn, Gs, Tr, tree height, ground diameter, holocellulose content and fiber length, and reduce the lignin content of P. tomentosa. The results showed that GA could significantly affect the physiology, photosynthesis, growth and wood quality traits of P. tomentosa.2. By using RNA-seq, this study identified 1,268 GA-responsive genes. Pathway enrichment analysis revealed that 87 GA-responsive genes were related to cell wall, chloroplast division, cell division, cell expansion, and photosynthesis. Regulatory motif enrichment analysis revealed that 37 GA-responsive genes related to photosynthesis shared two essential GA-related cis-regulatory elements, the GA response element （GARE） and the pyrimidine box. Thus, based on the known GA signal transduction pathway, we constructed a GA-responsive pathway consisting of genes involved in regulating photosynthesis. SNP-based association mapping identified that 142 SNPs within 40 genes of the constructed pathway were significantly associated with photosynthetic, growth and wood property traits （Q< 0.10）, explaining 0.01%-20.89% of phenotypic variation. Epistasis analysis identified that 310 SNP-SNP pairs had epistatic effects on photosynthetic, growth and wood property traits, providing evidence for the interactions between genes in the photosynthetic GA response payhway.3. By using RNA-seq, this study identified 7,655 P. tomentosa lncRNAs at genome-wide. These lncRNAs are shorter and expressed at significantly lower levels than protein-coding transcripts. In addition, P. tomentosa lncRNAs exhibit a low degree of evolutionary constraint. Among the 7655 lncRNAs, the levels of 410 lncRNAs changed in response to GA. Computational analysis predicted 939 potential cis-regulated target genes and 965 potential trans-regulated target genes for GA-responsive lncRNAs. Functional annotation and pathway enrichment of these potential target genes showed that they may be involved in tree growth and wood formation. SNP-based association analysis showed that 38 SNPs from 20 GA-responsive lncRNA genes and 363 SNPs from 158 potential target genes were significantly associated with growth and wood properties traits （Q< 0.10）, explaining 2.50%-5.40% of phenotypic variation. Epistasis analysis uncovered interactions between 312 SNP-SNP pairs representing interactions between SNPs within lncRNAs and potential target genes, providing evidence for interactions between lncRNAs and their potential target genes.4. By using miRNA-seq, this study identified 292 known miRNAs and 103 novel miRNAs in P. tomentosa at genome-wide. Among these,25 known miRNAs and 33 novel miRNAs were GA-responsive. Target gene prediction identified that these 58 GA-responsive miRNAs may interact with 345 potential target genes. Functional annotation and pathway enrichment of these potential target genes showed that they may be related to growth. SNP diversity analysis showed that the mature region of miRNAs and target region of target genes were highly conservative. SNP-based association analysis showed that 38 SNPs from 27 GA-responsive miRNAs and 176 SNPs from 87 potential target genes were significantly associated with photosynthetic and growth traits （Q< 0.10）, explaining 2.97%-7.13% of phenotypic variation. The association results suggested that these SNPs may have influence on photosynthetic and growth traits. Epistasis analysis uncovered interactions between 82 SNP-SNP pairs representing interactions between SNPs within miRNAs and potential target genes, providing evidence for interactions between miRNAs and their potential target genes.5. Analyses of the interactions among genes, lncRNAs and miRNAs revealed that 10 lncRNAs were predicted to be the precursors of 17 known miRNAs which may interact with 167 target genes, and 192 lncRNAs were predicted to be the targets of 191 known miRNAs. In addition, TCONS₀0264314 were predicted to be target mimic of ptc-miR6459b while TCONS₀0015177 and TCONS₀0013311 were predicted to be target mimic of ptc-miR169y. This study also identified complex interactions among genes, lncRNAs and miRNAs in the auxin signal transduction pathway, indicating that lncRNAs and miRNAs may be involved in the signal transduction of auxin.