Wuschel overexpression promotes somatic embryogenesis and induces organogenesis in cotton (Gossypium hirsutum L.) tissues cultured in vitro

• DOI: 10.1007/s00299-013-1402-9
• Publication Date: 2013-03-29T06:35:32Z
• Authors: O. Bouchabké-Coussa; M. Obellianne; D. Linderme; E. Montes; A. Maia-Grondard; F. Vilaine; C. Pannetier
• Keywords: EXPRESSION; [SDV.SA]Life Sciences [q-bio]/Agricultural sciences; 0301 basic medicine; 570; ARABIDOPSIS SHOOT MERISTEM; Organogenesis; Recombinant Fusion Proteins; Green Fluorescent Proteins; Molecular Sequence Data; Gossypium hirsutum; F62 - Physiologie végétale - Croissance et développement; PLANT STEM-CELLS; Tissue Culture Techniques; 03 medical and health sciences; REGENERATION; Gene Expression Regulation, Plant; WUSCHEL; AGROBACTERIUM-MEDIATED TRANSFORMATION; 580; Homeodomain Proteins; EMBRYO DEVELOPMENT; Gossypium; CAUSES ECTOPIC MORPHOGENESIS; Base Sequence; Indoleacetic Acids; Arabidopsis Proteins; http://aims.fao.org/aos/agrovoc/c_3339; Somatic embryogenesis; Plants, Genetically Modified; GENE; MAINTENANCE; F02 - Multiplication végétative des plantes; Agrobacterium tumefaciens; Seeds; Transformation, Bacterial; FLORAL MERISTEMS

This work shows that overexpression of the WUS gene from Arabidopsis enhanced the expression of embryogenic competence and triggered organogenesis from some cells of the regenerated embryo-like structures. Agrobacterium-mediated genetic transformation of cotton was described in the late 1980s, but is still time consuming and largely genotype dependant due to poor regeneration. To help solve this bottleneck, we over-expressed the WUSCHEL (WUS) gene, a homeobox transcription factor cloned in Arabidopsis thaliana, known to stimulate organogenesis and/or somatic embryogenesis in Arabidopsis tissues cultured in vitro. The AtWUS gene alone, and AtWUS gene fused to the GFP marker were compared to the GFP gene alone and to an empty construct used as a control. Somatic embryogenesis was improved in WUS expressed calli, as the percentage of explants giving rise to embryogenic tissues was significantly higher (×3) when WUS gene was over-expressed than in the control. An interesting result was that WUS embryogenic lines evolved in green embryo-like structures giving rise to ectopic organogenesis never observed in any of our previous transformation experiments. Using our standard in vitro culture protocol, the overexpression of AtWUS in tissues of a recalcitrant variety did not result in the production of regenerated plants. This achievement will still require the optimization of other non-genetic factors, such as the balance of exogenous phytohormones. However, our results suggest that targeted expression of the WUS gene is a promising strategy to improve gene transfer in recalcitrant cotton cultivars.