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Background  The frequency of cyanobacterial blooms has increased worldwide, and these blooms have been claimed to be a major factor leading to the decline of the most important freshwater herbivores, i.e. representatives of the genus  Daphnia . This suppression of  Daphnia  is partly attributed to the presence of biologically active secondary metabolites in cyanobacteria. Among these metabolites, protease inhibitors are found in almost every natural cyanobacterial bloom and have been shown to specifically inhibit  Daphnia 's digestive proteases  in vitro , but to date no physiological responses of these serine proteases to cyanobacterial protease inhibitors in  Daphnia  have been reported  in situ  at the protein and genetic levels.    Results  Nine digestive proteases were detected in  D. magna  using activity-stained SDS-PAGE. Subsequent analyses by LC-MS/MS and database search led to the identification of respective protease genes.  D. magna  responded to dietary protease inhibitors by up-regulation of the expression of these respective proteases at the RNA-level and by the induction of new and less sensitive protease isoforms at the protein level. The up-regulation in response to dietary trypsin- and chymotrypsin-inhibitors ranged from 1.4-fold to 25.6-fold. These physiological responses of  Daphnia , i.e. up-regulation of protease expression and the induction of isoforms, took place even after feeding on 20% cyanobacterial food for only 24 h. These physiological responses proved to be independent from microcystin effects.    Conclusion  Here for the first time it was shown  in situ  that a  D. magna  clone responds physiologically to dietary cyanobacterial protease inhibitors by phenotypic plasticity of the targets of these specific inhibitors, i.e.  Daphnia  gut proteases. These regulatory responses are adaptive for  D. magna , as they increase the capacity for protein digestion in the presence of dietary protease inhibitors. The type and extent of these responses in protease expression might determine the degree of growth reduction in  D. magna  in the presence of cyanobacterial protease inhibitors. The rapid response of  Daphnia  to cyanobacterial protease inhibitors supports the assumption that dietary cyanobacterial protease inhibitors exert a strong selection pressure on  Daphnia  proteases themselves.

Gene expression and activity of digestive proteases in Daphnia: effects of cyanobacterial protease inhibitors