Inducible tolerance to dietary protease inhibitors in Daphnia magna

Abstract

Daphnia has been shown to acquire tolerance to cyanobacterial toxins within an animals' lifetime and to transfer this tolerance to the next generation. Here we used a strain of the cyanobacterium Microcystis aeruginosa, which contained two chymotrypsin inhibitors (BN920 and CP954), the green alga Scenedesmus obliquus as reference food and a clone of D. magna to investigate the physiological mechanism of acquired tolerance to these cyanobacterial toxins. The intracellular concentrations of CP954 and BN920 were 1550 and 120 μmol l–1, respectively. When food suspensions of the green alga contained >60% M. aeruginosa, growth rates of D. magna were reduced. When grown on the green alga, three chymotrypsins ranging in mass from 16 to 22 kDa were distinguished in D. magna. Purified BN920 and CP954 specifically inhibited D. magna chymotrypsins. Feeding with encapsulated BN920 resulted in growth depression in D. magna and replacement of the chymotrypsins by three chymotrypsins with smaller molecular mass. With just 20% M. aeruginosa, the same changes in the chymotrypsin pattern as with the pure inhibitor were observed. IC50 values for inhibition of chymotrypsins of D. magna growing on the green alga were 5.4 nmol l–1 (BN920) and 7.4 nmol l–1 (CP954). When D. magna was grown on 20% M. aeruginosa, 2.2-fold higher IC50 values were observed. This indicated that increased tolerance to these dietary inhibitors was acquired within an animal's lifetime by remodelling the digestive chymotrypsins, which in turn serves as an intra-generational defence against these cyanobacterial inhibitors. This mechanism might be relevant for the transfer of tolerance to the next generation through maternal effects.