Functional genomics analysis of foliar condensed tannin and phenolic glycoside regulation in natural cottonwood hybrids

Regulation of leaf condensed tannins (CT) and salicylate-derived phenolic glycosides (PG) in fast- and slow-growing cottonwood backcrosses was analyzed by metabolic profiling and cDNA microarray hybridization. Seven hybrid lines of Populus fremontii L. and P. angustifolia James exhibiting growth/CT-PG phenotypes ranging from fast/low (Lines 18 and 1979) to slow/high (Lines 1012 and RL2) and intermediate (Lines NUL, 3200 and RM5) were investigated. Methanol-extractable leaf metabolites were analyzed by gas chromatography—mass spectrometry, and the results evaluated by principal component analysis. The hybrid lines formed separate clusters based on their primary metabolite profiles, with cluster arrangement also reflecting differences in CT-PG phenotype. Nitrogen (N) supply was manipulated to alter CT-PG partitioning and to obtain molecular insights into how primary metabolism interfaces with CT-PG accumulation. Three backcross lines (RM5, 1012, 18) exhibiting differential CT-PG responses to a 10-day hydroponic N-deprivation treatment were chosen for metabolite and gene expression analyses. The fast- growing Line 18 showed a minimal CT-PG response to N deprivation, and a reduction in photosynthetic gene expression. Line 1012 exhibited a strong phenylpropanoid response to N deprivation, including a doubling in phenylalanine ammonia-lyase (PAL) gene expression, and a shift from CT accumulation in the absence of stress toward PG accumulation under N-deprivation conditions. Amino acid concentrations were depressed in Lines 18 and 1012, as was expression of nitrate-sensitive genes coding for transketolase (TK), and malate dehydrogenase (MDH). Genes associated with protein synthesis and fate were down-regulated in Line 1012 but not in Line 18. Line RM5 exhibited a comparatively large increase in CT in response to N deprivation, but did not sustain decreases in amino acid concentrations, or changes in PAL, TK or MDH gene expression. Molecular characterization of the variable CT-PG responses shows promise for the identification and future testing of candidate genes for CT-PG trait selection or manipulation.