Arabidopsis Group IIId ERF proteins positively regulate primary cell wall-type CESA genes.

The cell wall determines morphology and the environmental responses of plant cells. The primary cell wall (PCW) is produced during cell division and expansion, determining the cell shape and volume. After cell expansion, specific types of plant cells produce a lignified wall, known as a secondary cell wall (SCW). We functionally analyzed Group IIId Arabidopsis AP2/EREBP genes, namely ERF34, ERF35, ERF38, and ERF39, which are homologs of a rice ERF gene previously proposed to be related to SCW biosynthesis. Expression analysis revealed that these four genes are expressed in regions related to cell division and/or cell differentiation in seedlings (i.e., shoot apical meristems, the primordia of leaves and lateral roots, trichomes, and central cylinder of primary roots) and flowers (i.e., vascular tissues of floral organs and replums and/or valve margins of pistils). Overexpression of ERF genes significantly upregulated PCW-type, but not SCW-type, CESA genes encoding cellulose synthase catalytic subunits in Arabidopsis seedlings. Transient co-expression reporter analysis indicated that ERF35, ERF38, and ERF39 possess transcriptional activator activity, and that ERF34, ERF35, ERF38, and ERF39 upregulated the promoter activity of CESA1, a PCW-type CESA gene, through the DRECRTCOREAT elements, the core cis-acting elements known to be recognized by AP2/ERF proteins. Together, our findings show that Group IIId ERF genes are positive transcriptional regulators of PCW-type CESA genes in Arabidopsis and are possibly involved in modulating cellulose biosynthesis in response to developmental requirements and environmental stimuli.

Evaluation of a morphological marker selection and excision system to generate marker-free transgenic cassava plants.

The efficacy of the ipt-type Multi-Auto-Transformation (MAT) vector system to transform the extensively grown cassava cultivar "KU50" was evaluated. This system utilizes the isopentenyltransferase (ipt) gene as morphological marker for visual selection of transgenic lines. The extreme shooty phenotype (ESP) of transgenic lines is lost due to the removal of ipt gene mediated by the yeast Rint/RS system. As a result, phenotypically normal shoots, considered marker-free transgenic plants, could be obtained. When transforming KU50 cassava cultivar with two different ipt-type MAT vectors, transformation frequency at 19-21% was observed. Among the total number of ESP explants, 32-38% regained normal extended shoot phenotype and 88-96% of which were confirmed to represent the marker-free transgenic plants. This is the first demonstration of the efficacy of Rint/RS system in promoting excision of ipt marker gene in cassava specie, with the consequent rapid production of marker-free transgenic plants. The high efficiency of this system should facilitate pyramiding a number of transgenes by repeated transformation without having to undergo through laborious, expensive and time-consuming processes of sexual crossing and seed production. The generation of marker-free, thus environmentally safe, genetically modified cassava clones should also ease the public concerns regarding the use of transgenic cassava in both food and nonfood industries.