Interplay of calcium, cAMP and PKA in flavonoid accumulation by cell cultures of Hypericum androsaemum L.

Abstract

Widely distributed throughout Europe, Hypericum androsaemum L. is an herbaceous plant whose leaves have been used traditionally to prepare infusions with diuretic, cholagogue, and hepatoprotective properties. Recent research has also revealed that plant water extracts exhibit antitumor [1], antioxidant and anti-acetylcholinesterase activities [2]. Cell cultures established from hypocotyl-derived callus of H. androsaemum were reported [3] to accumulate small amounts of flavonoids, with the highest levels being achieved during the stationary phase (day 14). Later on [4], it was found that flavonoid accumulation and phenylalanine ammonia-lyase (PAL) activity increased considerably 72h after addition of CaCl2 (15 mM) or calcium ionophore A23187 (5µM) to the nutrient medium of 11-day-old cultures. Since there is a growing body of evidence that Ca2+ and cAMP interact in many systems, similar experiments were carried out using 100µM dibutyryl cAMP (db-cAMP), a membrane-permeable cAMP analog. Treatment with this cAMP-modulator also caused a pronounced increase in both flavonoid content and PAL activity measured on day 14. Moreover, the stimulatory effects of db-cAMP were prevented or markedly inhibited by pretreatment of cells with the calcium channel blocker verapamil (100µM). On the other hand, the Ca2+-induced enhancement of flavonoid accumulation and PAL activity could be mimicked by treating cell cultures with 50µM Sp-cAMPS, an activator of cAMP-dependent protein kinase A (PKA). Altogether, these data are consistent with a possible interplay between Ca2+, cAMP and PKA in the regulation of flavonoid accumulation in H. androsaemum cell cultures.