title: SND1 transcription factor-directed quantitative functional hierarchical genetic regulatory network in wood formation in Populus trichocarpa.
abstract: Wood is an essential renewable raw material for industrial products and energy. However, knowledge of the genetic regulation of wood formation is limited. We developed a genome-wide high-throughput system for the discovery and validation of specific transcription factor (TF)-directed hierarchical gene regulatory networks (hGRNs) in wood formation. This system depends on a new robust procedure for isolation and transfection of Populus trichocarpa stem differentiating xylem protoplasts. We overexpressed Secondary Wall-Associated NAC Domain 1s (Ptr-SND1-B1), a TF gene affecting wood formation, in these protoplasts and identified differentially expressed genes by RNA sequencing. Direct Ptr-SND1-B1-DNA interactions were then inferred by integration of time-course RNA sequencing data and top-down Graphical Gaussian Modeling-based algorithms. These Ptr-SND1-B1-DNA interactions were verified to function in differentiating xylem by anti-PtrSND1-B1 antibody-based chromatin immunoprecipitation (97% accuracy) and in stable transgenic P. trichocarpa (90% accuracy). In this way, we established a Ptr-SND1-B1-directed quantitative hGRN involving 76 direct targets, including eight TF and 61 enzyme-coding genes previously unidentified as targets. The network can be extended to the third layer from the second-layer TFs by computation or by overexpression of a second-layer TF to identify a new group of direct targets (third layer). This approach would allow the sequential establishment, one two-layered hGRN at a time, of all layers involved in a more comprehensive hGRN. Our approach may be particularly useful to study hGRNs in complex processes in plant species resistant to stable genetic transformation and where mutants are unavailable.
authors.affiliation: Forest Biotechnology Group, Department of Forestry and Environmental Resources, North Carolina State University, Raleigh, North Carolina 27695.
authors.forename: Ying-Chung
authors.initials: YC
authors.lastname: Lin
authors.forename: Wei
authors.initials: W
authors.lastname: Li
authors.forename: Ying-Hsuan
authors.initials: YH
authors.lastname: Sun
authors.forename: Sapna
authors.initials: S
authors.lastname: Kumari
authors.forename: Hairong
authors.initials: H
authors.lastname: Wei
authors.forename: Quanzi
authors.initials: Q
authors.lastname: Li
authors.forename: Sermsawat
authors.initials: S
authors.lastname: Tunlaya-Anukit
authors.forename: Ronald R
authors.initials: RR
authors.lastname: Sederoff
authors.forename: Vincent L
authors.initials: VL
authors.lastname: Chiang
mesh_terms: D000465:Algorithms; D002473:Cell Wall / Q000235:genetics; D047369:Chromatin Immunoprecipitation; D018506:Gene Expression Regulation, Plant; D063990:Gene Ontology; D053263:Gene Regulatory Networks; D059014:High-Throughput Nucleotide Sequencing; D008957:Models, Genetic; D010940:Plant Proteins / Q000235:genetics* / Q000378:metabolism; D030821:Plants, Genetically Modified; D016133:Polymerase Chain Reaction / Q000379:methods; D032107:Populus / Q000235:genetics* / Q000378:metabolism; D011523:Protoplasts; D015203:Reproducibility of Results; D014157:Transcription Factors / Q000235:genetics / Q000378:metabolism*; D014162:Transfection; D014934:Wood / Q000235:genetics* / Q000378:metabolism; D052584:Xylem / Q000166:cytology / Q000235:genetics
chemical_list: D010940:Plant Proteins; D014157:Transcription Factors