Type II Metacaspases are involved in cell stress but not in cell death in the unicellular green alga Dunaliella tertiolecta.

Abstract

Ultraviolet radiation (UVR; 280–400 nm) has a great impact on aquatic ecosystems by affecting ecophysiological and biogeochemical pro-cesses as a consequence of the global change scenario generated by anthro-pogenic activities. We studied the effect of PAR (P)+UVA (A)+UVB (B) i.e. PAB, on the molecular physiology of the unicellular green alga Dunaliella tertio-lecta for six days. We assessed the relationship between the triggered UVR stress response and metacaspases and caspase-like (CL)activities, which are proteases denoted to participate in cell death (CD) in phytoplankton. UVR inhibited cell growth and in vivo chlorophyll a fluorescence but did not cause cell death. Western blot analyses reflected that Type-II metacaspases (MCs) are present and appear to be involved in UVR induced-cell stress but not in dark-induced CD in D. tertiolecta. Enzyme kinetics revealed that cleavage of the MCs-reporter substrates RVRR, QRR, GRR, LKR, HEK, and VLK was 10-fold higher than WEHD, DEVD, IETD, and LETD CLs-substrates. The lowest apparent Michaelis-Menten constants (KMap) corresponded to RVRRase (37.5 μM) indi-cating a high affinity by the RVRR substrate. The inhibition of enzymatic activ-ities by using inhibitors with different target sites for hydrolyses demonstrat-ed that from all of the R/ Kase activities only RVRRase was a potential candi-date for being a metacaspase. In parallel, zymograms and peptide-mass fin-gerprinting analyses revealed the identities of such Rase activities suggesting an indirect evidence of possible natural physiological substrates of MCs. We present evidence of type II-MCs not being involved in CD in D. tertiolecta, but rather in survival strategies under the stressful irradiance conditions applied in this study.