title: Incipient stem cell niche conversion in tissue culture: using a systems approach to probe early events in WUSCHEL-dependent conversion of lateral root primordia into shoot meristems.
abstract: Adventitious shoot organogenesis contributes to the fitness of diverse plant species, and control of this process is a vital step in plant transformation and in vitro propagation. New shoot meristems (SMs) can be induced by the conversion of lateral root primorida/meristems (LRP/LRMs) or callus expressing markers for this identity. To study this important and fascinating process we developed a high-throughput methodology for the synchronous initiation of LRP by auxin, and subsequent cytokinin-induced conversion of these LRP to SMs. Cytokinin treatment induces the expression of the shoot meristematic gene WUSCHEL (WUS) in converting LRP (cLRP) within 24-30 h, and WUS is required for LRP → SM conversion. Subsequently, a transcriptional reporter for CLAVATA3 (CLV3) appeared 32-48 h after transfer to cytokinin, marking presumptive shoot stem cells at the apex of cLRP. Thus the spatial expression of these two components (WUS and CLV3) of a regulatory network maintaining SM stem cells already resembles that seen in a vegetative shoot apical meristem (SAM), suggesting the very rapid initiation and establishment of the new SMs. Our high-throughput methodology enabled us to successfully apply a systems approach to the study of plant regeneration. Herein we characterize transcriptional reporter expression and global gene expression changes during LRP → SM conversion, elaborate the role of WUS and WUS-responsive genes in the conversion process, identify and test putative functional targets, perform a comparative analysis of domain-specific expression in cLRP and SM tissue, and develop a bioinformatic tool for examining gene expression in diverse regeneration systems.
authors.affiliation: Department of Cell and Systems Biology, University of Toronto, 25 Harbord Street, Toronto, ON M5S 3G5, Canada. steven_Chatfield@carleton.ca
authors.forename: Steven P
authors.initials: SP
authors.lastname: Chatfield
authors.forename: Resmi
authors.initials: R
authors.lastname: Capron
authors.forename: Andre
authors.initials: A
authors.lastname: Severino
authors.forename: Pier-Andree
authors.initials: PA
authors.lastname: Penttila
authors.forename: Simon
authors.initials: S
authors.lastname: Alfred
authors.forename: Hardeep
authors.initials: H
authors.lastname: Nahal
authors.forename: Nicholas J
authors.initials: NJ
authors.lastname: Provart
mesh_terms: D017360:Arabidopsis / Q000166:cytology / Q000235:genetics / Q000502:physiology*; D029681:Arabidopsis Proteins / Q000235:genetics* / Q000378:metabolism; D002454:Cell Differentiation; D003583:Cytokinins / Q000378:metabolism; D015536:Down-Regulation; D020869:Gene Expression Profiling; D018507:Gene Expression Regulation, Developmental; D018506:Gene Expression Regulation, Plant; D055786:Gene Knockout Techniques; D017930:Genes, Reporter; D005838:Genotype; D018398:Homeodomain Proteins / Q000235:genetics* / Q000378:metabolism; D007210:Indoleacetic Acids / Q000378:metabolism; D018519:Meristem / Q000166:cytology / Q000235:genetics / Q000502:physiology; D020411:Oligonucleotide Array Sequence Analysis; D010937:Plant Growth Regulators / Q000378:metabolism*; D018517:Plant Roots / Q000166:cytology / Q000235:genetics / Q000502:physiology; D018520:Plant Shoots / Q000166:cytology / Q000235:genetics / Q000502:physiology; D030821:Plants, Genetically Modified; D011993:Recombinant Fusion Proteins; D012038:Regeneration; D055153:Stem Cell Niche / Q000502:physiology*; D046509:Tissue Culture Techniques; D059467:Transcriptome; D015854:Up-Regulation
chemical_list: C118167:AT2G27250 protein, Arabidopsis; D029681:Arabidopsis Proteins; D003583:Cytokinins; D018398:Homeodomain Proteins; D007210:Indoleacetic Acids; D010937:Plant Growth Regulators; D011993:Recombinant Fusion Proteins; C116421:WUSCHEL protein, Arabidopsis