Effects of food deprivation on the respiratory metabolism of Oxyrrhismarina

Effects of food deprivation on the respiratory metabolism of Oxyrrhis marina N. Osmaa,*, T. Packarda, M. Gómeza aBiological Oceanography Laboratory, Facultad de Ciencias del Mar, Universidad de las Palmas de Gran Canaria, Campus Universitario de Tafira. 35017. Las Palmas de Gran Canaria. Canary Islands, Spain. * E–mail: [email protected] INTRODUCTION The microheterotroph Oxyrrhis marina has been used extensively as a model organism to examine the responses of heterotrophic flagellates to a variety of factors. Here, we focused in the respiratory metabolism. Respiration is the oxygen consumption (RO2) catalyzed by the enzymes of the respiratory electron transfer system (ETS). The ETS transfers the electrons from the reducing equivalents that are produced during the oxidation of organic molecules to oxygen (Fig 1). Thus, the RO2 rate of an organism is related to the food availability. We fed O. marina with the cryptophyta Rhodomonas salina and we elucidated the variation of the respiration rates in starvation. METHODS & RESULTS CONCLUSIONS I) Respiration decreases during food deprivation in O. marina. II) ETS activity is constant during food deprivation in O. marina. III) Cell sizes increases during food deprivation in O. marina (cannibalism?). IV) The R/ETS ratio decreases both during food deprivation and with increasing cell size in O. marina. Aknowledgements This research, is funded by the Formation and Perfection of the Researcher Personal Program of the Basque Country Government and by the EXZOME project (CTM 2008 –01616/MAR) from the Spanish Science and Education Ministry. References: Lowry, O. H., N. J. Rosebrough, et al. (1951). "Protein measurement with the folin phenol reagent." Journal of Biological Chemistry 193: 265 - 275. Packard, T. T., E. Berdalet, et al. (1996). "Oxygen consumption in the marine bacterium Pseudomonas nautica predicted from ETS activity and bisubstrate enzyme kinetics." Journal of Plankton Research 18(10): 1819 –1835. Figure 1. Structure of the Electron Transfer System (ETS) coupled with energy (ATP) formation. START SAMPLING!! END SAMPLING!! 0 20 40 60 80 100 120 140 1 2 3 4 RHODOMONAS (Cell number x106/l) 0 5 10 15 20 25 30 35 0 20 40 60 80 100 120 140 1 3 5 7 9 11 13 15 17 PROTEIN (mgl-1) or CELL NUMBER (x106/l) RO2or ETS (nmol O2min-1l-1) I II III IV 1 3 5 7 9 11 13 15 17 TIME(days) PROTEIN ETS CELL NUMBER RESPIRATION Figure 2. (A) Cell number decline of Rhodomonas salina in the culture due to the feeding by O. marina. The experiment began when the algae concentration in the culture was negligible. (B) Time profiles of the measurements taken every 2 days of protein, ETS, respiration and cell number of O. marina.The protein content and the ETS activity stayed stable during the entire experiment while the respiration and the cell number decreased more than 50 %in the first nine days. Each measurement represents the mean of triplicate analyses (n =27). AB FEEDING!! Figure 3. Time profiles of: (A) specific respiration; (B) ETS activity; and (C) protein content per cell of O. marina. The consumption of O2per cell decreased during the experiment while the ETS activity and protein content increased. Each measurement represents the mean of triplicate analyses (n=27). y = -0,0121x + 0,257 R² = 0,7623 0,0 0,1 0,2 0,3 0,4 0246810 12 14 16 18 RO2 (pmol O2h-1cell-1) TIME (days) y = 0,0407x + 0,102 R² = 0,7863 0,0 0,2 0,4 0,6 0,8 1,0 0 2 4 6 8 10 12 14 16 18 ETS (pmol O2h-1cell-1) TIME (days) y = 0,2105x + 1,2298 R² = 0,8857 0 1 2 3 4 5 6 7 8 0 2 4 6 8 10 12 14 16 18 Protein (ng cell-1) TIME (days) y = -0,589ln(x) + 1,7285 R² = 0,9214 0 0,5 1 1,5 2 0246810 12 14 16 18 R/ETS TIME (Days) y = -1,194ln(x) + 1,8581 R² = 0,8795 0 0,5 1 1,5 2 1 2 3 4 5 R/ETS Protein/cell (ng/cell) Figure 4. (A) Time course of the respiration / ETS activity ratio during the experiment. (B) Variation of the R/ETS ratio with cell specific protein (cell size). The R/ETS ratio declined exponentially as long as O. marina was starved (left). The same behaviour in the ratio occurred as the cell size increased (right). The data show a Pearson correlation coefficient of -0,902 (n=9, p<0,01). BA ACB Oxyrrhis marina. Rhodomonas salina. ALGAL CULTURING Triplicate cultures of O. marina,at 22.5ºC and low irradiance. Filtered Samples ETS activity Packard et al (1996) *adding succinate Protein Lowry et al. (1951) In vivo Samples Cell number Lugol (4%), Counting chamber Respiration Oxygen electrodes. ANALYTICAL METHODS MITOCHONDRIA