Ubiquity and quantitative significance of detoxification catabolism of chlorophyll associated with protistan herbivory

Abstract

Chlorophylls are essential components of the photosynthetic apparati that sustain all of the life forms that ultimately depend on solar energy. However, a drawback of the extraordinary photosensitizing efficiency of certain chlorophyll species is their ability to generate harmful singlet oxygen. Recent studies have clarified the catabolic processes involved in the detoxification of chlorophylls in land plants, but little is understood about these strategies in aquatic ecosystem. Here, we report that a variety of heterotrophic protists accumulate the chlorophyll a catabolite 13 2 ,17 3 -cyclopheophorbide a enol (cPPB-aE) after their ingestion of algae. This chlorophyll derivative is nonfluorescent in solution, and its inability to generate singlet oxygen in vitro qualifies it as a detoxified catabolite of chlorophyll a. Using a modified analytical method, we show that cPPBaE is ubiquitous in aquatic environments, and it is often the major chlorophyll a derivative. Our findings suggest that cPPB-aE metabolism is one of the most important, widely distributed processes in aquatic ecosystems. Therefore, the herbivorous protists that convert chlorophyll a to cPPB-aE are suggested to play more significant roles in the modern oceanic carbon flux than was previously recognized, critically linking microscopic primary producers to the macroscopic food web and carbon sequestration in the ocean. phototoxicity of chlorophylls | microbial herbivory | phagocytosis | biodiversity of eukaryotes | microbial loop C hlorophylls are crucial to sustaining most life forms on Earth, the majority of which ultimately depend on solar energy. Photoexcitation of chlorophylls initiates various photosynthetic reactions that convert the energy of photons into chemical potentials, which in turn, drive the full range of metabolic reactions throughout the global ecosystem. Chlorophylls play a central role in the photosynthetic apparatus by absorbing light and transferring the excitation energy to the reaction centers of photosystems before photosynthetic electron transport. However, without measures to contain the excited energy, chlorophylls can harm organisms because of their high photosensitizing potential.

Risk Management by Organisms of the Phototoxicity of Chlorophylls

“…37 We cultured a series of phycophagic protists and a specific unicellular alga as their food, and in an analysis of the pigment extracts of the cultures, we found CPEs to be the sole major chlorophyll derivatives. 35,36 Significantly, we found that CPEs are essentially nonfluorescent in solution (Φ F < 0.002 for cPPB-aE). 35 Under visible-light illumination, cPPB-aE in air-saturated anisole generated no 1 O 2 , whereas parallel experiments with Chl-a and Phe-a rapidly produced 1 O 2 .…”

“…35,36 Significantly, we found that CPEs are essentially nonfluorescent in solution (Φ F < 0.002 for cPPB-aE). 35 Under visible-light illumination, cPPB-aE in air-saturated anisole generated no 1 O 2 , whereas parallel experiments with Chl-a and Phe-a rapidly produced 1 O 2 . 35 The photoexcited S 1 states of the CPEs were immediately quenched to S 0 by internal conversion (È S ic 1), resulting in little chance of intersystem crossing to their T 1 states (Φ T µ 0).…”

“…We consider that CPEs are the detoxified catabolites of chlorophylls, associated with the digestive processes of protists (phagocytosis). 35,36 In contrast to the processes in the PAO catabolism displayed by land plants, CPE catabolism seems to be a rather rapid process. We microscopically demonstrated the fast disappearance of chlorophyll autofluorescence in the chloroplasts of ingested algae in an early stage of phagocytosis by a protistan predator (within 10 min), suggesting the very rapid conversion of chlorophylls into nonradiatively quenching catabolites (Figure 3).…”

“…35 Under visible-light illumination, cPPB-aE in air-saturated anisole generated no 1 O 2 , whereas parallel experiments with Chl-a and Phe-a rapidly produced 1 O 2 . 35 The photoexcited S 1 states of the CPEs were immediately quenched to S 0 by internal conversion (È S ic 1), resulting in little chance of intersystem crossing to their T 1 states (Φ T µ 0). The quenching mechanism has not yet been clarified, but is probably attributable to their specific molecular structures, in which the β-diketonate moiety is π-conjugated with the tetrapyrrole macrocycle.…”

“…35,36 The derivatives have an exo-seven-membered ring formed by the Claisen-type condensation of the peripheral functional groups in the chlorin π-system of Chls-a/b: 13-COCH and 17-CH 2 CH 2 COO-phytyl. 37 We cultured a series of phycophagic protists and a specific unicellular alga as their food, and in an analysis of the pigment extracts of the cultures, we found CPEs to be the sole major chlorophyll derivatives.…”