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1.  Acclimatory responses of the Daphnia pulex proteome to environmental changes. II. Chronic exposure to different temperatures (10 and 20°C) mainly affects protein metabolism 
BMC Physiology  2009;9:8.
Temperature affects essentially every aspect of the biology of poikilothermic animals including the energy and mass budgets, activity, growth, and reproduction. While thermal effects in ecologically important groups such as daphnids have been intensively studied at the ecosystem level and at least partly at the organismic level, much less is known about the molecular mechanisms underlying the acclimation to different temperatures. By using 2D gel electrophoresis and mass spectrometry, the present study identified the major elements of the temperature-induced subset of the proteome from differently acclimated Daphnia pulex.
Specific sets of proteins were found to be differentially expressed in 10°C or 20°C acclimated D. pulex. Most cold-repressed proteins comprised secretory enzymes which are involved in protein digestion (trypsins, chymotrypsins, astacin, carboxypeptidases). The cold-induced sets of proteins included several vitellogenin and actin isoforms (cytoplasmic and muscle-specific), and an AAA+ ATPase. Carbohydrate-modifying enzymes were constitutively expressed or down-regulated in the cold.
Specific sets of cold-repressed and cold-induced proteins in D. pulex can be related to changes in the cellular demand for amino acids or to the compensatory control of physiological processes. The increase of proteolytic enzyme concentration and the decrease of vitellogenin, actin and total protein concentration between 10°C and 20°C acclimated animals reflect the increased amino-acids demand and the reduced protein reserves in the animal's body. Conversely, the increase of actin concentration in cold-acclimated animals may contribute to a compensatory mechanism which ensures the relative constancy of muscular performance. The sheer number of peptidase genes (serine-peptidase-like: > 200, astacin-like: 36, carboxypeptidase-like: 30) in the D. pulex genome suggests large-scaled gene family expansions that might reflect specific adaptations to the lifestyle of a planktonic filter feeder in a highly variable aquatic environment.
PMCID: PMC2678069  PMID: 19383147
2.  Acclimatory responses of the Daphnia pulex proteome to environmental changes. I. Chronic exposure to hypoxia affects the oxygen transport system and carbohydrate metabolism 
BMC Physiology  2009;9:7.
Freshwater planktonic crustaceans of the genus Daphnia show a remarkable plasticity to cope with environmental changes in oxygen concentration and temperature. One of the key proteins of adaptive gene control in Daphnia pulex under hypoxia is hemoglobin (Hb), which increases in hemolymph concentration by an order of magnitude and shows an enhanced oxygen affinity due to changes in subunit composition. To explore the full spectrum of adaptive protein expression in response to low-oxygen conditions, two-dimensional gel electrophoresis and mass spectrometry were used to analyze the proteome composition of animals acclimated to normoxia (oxygen partial pressure [Po2]: 20 kPa) and hypoxia (Po2: 3 kPa), respectively.
The comparative proteome analysis showed an up-regulation of more than 50 protein spots under hypoxia. Identification of a major share of these spots revealed acclimatory changes for Hb, glycolytic enzymes (enolase), and enzymes involved in the degradation of storage and structural carbohydrates (e.g. cellubiohydrolase). Proteolytic enzymes remained constitutively expressed on a high level.
Acclimatory adjustments of the D. pulex proteome to hypoxia included a strong induction of Hb and carbohydrate-degrading enzymes. The scenario of adaptive protein expression under environmental hypoxia can be interpreted as a process to improve oxygen transport and carbohydrate provision for the maintenance of ATP production, even during short episodes of tissue hypoxia requiring support from anaerobic metabolism.
PMCID: PMC2678976  PMID: 19383146
3.  Physiological responses of Daphnia pulex to acid stress 
BMC Physiology  2009;9:9.
Acidity exerts a determining influence on the composition and diversity of freshwater faunas. While the physiological implications of freshwater acidification have been intensively studied in teleost fish and crayfish, much less is known about the acid-stress physiology of ecologically important groups such as cladoceran zooplankton. This study analyzed the extracellular acid-base state and CO2 partial pressure (PCO2), circulation and ventilation, as well as the respiration rate of Daphnia pulex acclimated to acidic (pH 5.5 and 6.0) and circumneutral (pH 7.8) conditions.
D. pulex had a remarkably high extracellular pH of 8.33 and extracellular PCO2 of 0.56 kPa under normal ambient conditions (pH 7.8 and normocapnia). The hemolymph had a high bicarbonate concentration of 20.9 mM and a total buffer value of 51.5 meq L-1 pH-1. Bicarbonate covered 93% of the total buffer value. Acidic conditions induced a slight acidosis (ΔpH = 0.16–0.23), a 30–65% bicarbonate loss, and elevated systemic activities (tachycardia, hyperventilation, hypermetabolism). pH 6.0 animals partly compensated the bicarbonate loss by increasing the non-bicarbonate buffer value from 2.0 to 5.1 meq L-1 pH-1. The extracellular PCO2 of pH 5.5 animals was significantly reduced to 0.33 kPa, and these animals showed the highest tolerance to a short-term exposure to severe acid stress.
Chronic exposure to acidic conditions had a pervasive impact on Daphnia's physiology including acid-base balance, extracellular PCO2, circulation and ventilation, and energy metabolism. Compensatory changes in extracellular non-bicarbonate buffering capacity and the improved tolerance to severe acid stress indicated the activation of defense mechanisms which may result from gene-expression mediated adjustments in hemolymph buffer proteins and in epithelial properties. Mechanistic analyses of the interdependence between extracellular acid-base balance and CO2 transport raised the question of whether a carbonic anhydrase (CA) is involved in the catalysis of the reaction, which led to the discovery of 31 CA-genes in the genome of D. pulex.
PMCID: PMC2689847  PMID: 19383148

Results 1-3 (3)