Abstract
Seed dormancy release of the obligate root parasitic plant, Phelipanche ramosa, requires a minimum 4-day conditioning period followed by stimulation by host-derived germination stimulants, such as strigolactones. Germination is then mediated by germination stimulant-dependent activation of PrCYP707A1, an abscisic acid catabolic gene. The molecular mechanisms occurring during the conditioning period that silence PrCYP707A1 expression and regulate germination stimulant response are almost unknown. Here, global DNA methylation quantification associated with pharmacological approaches and cytosine methylation analysis of the PrCYP707A1 promoter were used to investigate the modulation and possible role of DNA methylation during the conditioning period and in the PrCYP707A1 response to GR24, a synthetic strigolactone analogue. Active global DNA demethylation occurs during the conditioning period and is required for PrCYP707A1 activation by GR24 and for subsequent seed germination. Treatment with 5-azacytidine, a DNA-hypomethylating molecule, reduces the length of the conditioning period. Conversely, hydroxyurea, a hypermethylating agent, inhibits PrCYP707A1 expression and seed germination. Methylated DNA immunoprecipitation followed by PCR experiments and bisulfite sequencing revealed that DNA demethylation particularly impacts a 78-nucleotide sequence in the PrCYP707A1 promoter. The results here demonstrate that the DNA methylation status during the conditioning period plays a crucial role independently of abscisic acid in the regulation of P. ramosa seed germination by controlling the strigolactone-dependent expression of PrCYP707A1.
Highlights
Some parasitic plants are deadly pests with the capacity to exploit other plants
Because abscisic acid (ABA) was shown to inhibit seed germination in P. ramosa (Pouvreau et al, 2013) and was suggested to control the acquisition of sensitivity to germination stimulants (GS) during the conditioning period (Lechat et al, 2012), seeds were conditioned in incubation medium (IM) supplemented or not with ABA for different durations in order to investigate the impact of the hormone on the capacity of seeds to respond to SL
When incubated in IM supplemented with 250 μM ABA and treated with 1 nM GR24, seeds did not germinate after 4 or 7 days of conditioning (5 ± 2% and 7 ± 5%), in contrast to seeds incubated in ABA-free IM (74 ± 6% and 86 ± 6%) (Fig. 1A)
Summary
Some parasitic plants are deadly pests with the capacity to exploit other plants. Among the obligate root parasitic weeds, the holoparasites, which are devoid of chlorophyll and unable to carry out photosynthesis, totally rely on a host for their water, mineral, and carbohydrate supplies. The underlying process of this trophic exploitation, governed by a fine-tuned molecular dialogue between both partners, is an extraordinary example of adaptive plant biology operated by these parasitic organisms in the course of evolution. Among the remarkable morphological and physiological adaptations that characterize this transition, the requirement for the seeds to perceive molecules produced by host roots to germinate is probably the most intriguing one. The germination of broomrape species is a two-step process corresponding first to a conditioning period, thought to be required for the acquisition of the sensitivity to germination stimulants (GS) (Joel et al, 1991), followed by the chemical stimulation of the germination itself that ends with the radicle protrusion (Joel and Bar, 2013)
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