Ammonium regulates embryogenic potential in Cucurbita pepo through pH-mediated changes in endogenous auxin and abscisic acid

Abstract

When supplied as the sole nitrogen source, ammonium can induce somatic embryogenesis in some plant species, including pumpkin. To get an insight into the involvement of phytohormones in this adaptive developmental response to nitrogen availability, the status of endogenous abscisic acid (ABA), indole-3-acetic acid (IAA), and their conjugates was analyzed in an ammonium-induced embryogenic pumpkin culture. Results were compared with endogenous hormone status in an embryogenic culture induced by exogenously supplied 2,4-dichlorophenoxyacetic acid (2,4-D). In the NH4 +-induced somatic embryogenic culture, the maximum concentration of ABA was measured after 21 days of cultivation, while in the 2,4-D-induced culture ABA peaked after 7 days of cultivation, which suggested a possible activation of different ABA-regulated stress responses. The NH4 +-induced embryogenic culture was also characterized by increased accumulation of endogenous IAA to the level similar or higher than the one induced by 2,4-D. A considerable decrease (threefold) in endogenous IAA content was observed after buffering the NH4 +-medium with MES or after nitrate re-supply. In contrast, buffering the NH4 +-medium increased endogenous ABA by five times. Significant ammonium- and pH-related changes were also detected in ascorbate and glutathione redox status, and discussed regarding the corresponding changes in cell size and mitotic activity in the cultures. Furthermore, a negative correlation between free IAA content and the activity of ascorbate peroxidase (EC 1.11.1.11) and auxin-sensitive peroxidase CpAPRX (EC 1.11.1.7) was observed. The results strongly suggest a mediating role of external pH in the regulation of endogenous phytohormones and the ascorbate–glutathione cycle during ammonium-induced acquisition of embryogenic competence in pumpkin.