Microarray analyses of gene expression during adventitious root development in Pinus contorta

Abstract

In order to investigate the gene expression pattern during adventitious root development, RNA of Pinus contorta hypocotyls, pulse-treated with the auxin indole-3-butyric acid and harvested at distinct developmental time points of root development, was hybridized to microarrays containing 2,178 cDNAs from Pinus taeda. Over the period of observation of root development, the transcript levels of 220 genes changed significantly. During the root initiation phase, genes involved in cell replication and cell wall weakening and a transcript encoding a PINHEAD/ZWILLE-like protein were up-regulated, while genes related to auxin transport, photosynthesis, and cell wall synthesis were down-regulated. In addition, there were changes in transcript abundance of genes related to water stress. During the root meristem formation phase the transcript abundances of genes involved in auxin transport, auxin responsive transcription, and cell wall synthesis, and of a gene encoding a B-box zinc finger-like protein, increased, while those encoding proteins involved in cell wall weakening decreased. Changes of transcript abundance of genes related to water stress during the root meristem formation and root formation phase indicate that the plant roots had become functional in water transport. Simultaneously, genes involved in auxin transport were up-regulated, while genes related to cell wall modification were down-regulated. Finally, during the root elongation phase down-regulation of transcripts encoding proteins involved in cell replication and stress occurred. Based on the observed changes in transcript abundances, we suggest hypotheses about the relative importance of various physiological processes during the auxin-induced development of roots in P. contorta.

Figure 1.

Auxin-induced adventitious root development in P. contorta. Four-week-old hypocotyl cuttings were treated with 1.23 mm IBA for 6 h. The consecutive development of adventitious roots was monitored at 3-d intervals, starting day 0 with untreated plants. A, Drawings illustrating the different stages of adventitious root development (adapted from Lindroth et al., 2001b, with permission from Kluwer Academic Publishers). B, Schematic drawings illustrating transverse sections of the hypocotyl during each stage in the development. The hypocotyl split after 3 d. The early stage of an adventitious root primordium (AP) is visible at day 6, and the adventitious root meristem (AM) is formed by day 9. Fully developed roots are formed by day 12, which then start to elongate (adapted from Lindroth et al., 2001a, with permission from Elsevier).

Figure 2.

Genes differentially expressed during various phases of adventitious root development in P. contorta. A, Venn diagram showing the number of genes differentially expressed during specific developmental phases of adventitious root development. B, The 220 genes differentially expressed during specific developmental phases of root development were separated into two groups according to whether they were significantly up-regulated or down-regulated.

Figure 3.

Hierarchical clustering of 220 genes differentially expressed during adventitious root development in P. contorta. The fold changes of 220 genes differentially expressed during root development were supplied to the program Genesis (developed by Alexander Sturn, Institute for Biomedical Engineering, Graz University of Technology, 2000–2002). A genetree was created using the average linkage clustering method (Pearson correlation).