Location: National Peanut Research LaboratoryTitle: WUS and STM-based reporter genes for studying meristem development in poplar
|BAO, Y. - OREGON STATE UNIVERSITY|
|DHARMAWARDHANA, P. - OREGON STATE UNIVERSITY|
|Arias De Ares, Renee|
|ALLEN, M.B. - OREGON STATE UNIVERSITY|
|MA, C. - OREGON STATE UNIVERSITY|
|STRAUSS, S.H. - OREGON STATE UNIVERSITY|
Submitted to: Plant Cell Reports
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 2/8/2009
Publication Date: 3/12/2009
Citation: Bao, Y., Dharmawardhana, P., Arias, R.S., Allen, M., Ma, C., Strauss, S. 2009. WUS and STM-based reporter genes for studying meristem development in poplar. Plant Cell Reports 28:947-962.
Interpretive Summary: Two of the main genes expressed in apical meristems in plants are WUSHEL (WUS) and SHOOTMERISTEMLESS (STM). In order to study stem cell formation in plants, after dedifferentiation, we isolated the promoters of WUS and STM, and linked them to the reporter gene for Glucuronidase (GUS). Poplar plants were transformed with those molecular constructs and gene expression was analyzed. In order to choose the proper alleles of this triploid plant (poplar) for the studies, significant bioinformatics and microarray analyses were performed.
Technical Abstract: We describe the development of a reporter system for monitoring meristem initiation in poplar using promoters of poplar homologs to the meristem-active regulatory genes WUSCHEL (WUS) and SHOOTMERISTEMLESS (STM). When ~3 kb of the 5’ flanking regions of close homologs were used to drive expression of the GUSPlus gene, 50–60% of the transgenic events showed expression in apical and axillary meristems. However, expression was also common in other organs, including in leaf veins (40 and 46% of WUS and STM transgenic events, respectively) and hydathodes (56% of WUS transgenic events). Histochemical GUS staining of explants during callogenesis and shoot regeneration using in vitro stems as explants showed that expression was detectable prior to visible shoot development, starting 3–15 days after explants were placed onto callus inducing medium. A minority of WUS and STM events also showed expression in the cambium, phloem, or xylem of regenerated, greenhouse grown plants undergoing secondary growth. Based on microarray gene expression data, a paralog of poplar WUS was detectably up-regulated during shoot initiation, but the other paralog was not. Both paralogs of poplar STM were downregulated threefold to sixfold during early callus initiation. We identified 15–35 copies of cytokinin response regulator binding motifs (ARR1AT) and one copy of the auxin response element (AuxRE) in both promoters. Several of the events recovered may be useful for studying the process of primary and secondary meristem development, including treatments intended to stimulate meristem development to promote clonal propagation and genetic transformation.