Differential regulation of hsp70 genes in the freshwater key species Gammarus pulex (Crustacea, Amphipoda) exposed to thermal stress: effects of latitude and ontogeny.

Temperature is one of the main abiotic factors influencing the distribution and abundance of organisms. In the Rhône River Valley, populations of the crustacean Gammarus pulex are distributed along a 5 °C thermal gradient from the North to the South of the valley. In this present work, we investigated the heat shock response of G. pulex according to latitudinal distribution (northern vs. southern populations) and ontogeny (adults vs. embryos from early stages). We isolated two isoforms (one constitutive hsc70 and one inducible hsp70) of heat shock proteins 70 (HSP70) and quantitatively compared their amounts of mRNA after heat shocks, using real-time PCR. Whereas the hsc70 (constitutive) gene did not vary between the two populations, a significant effect of the population was observed on the expression of the hsp70 (inducible) gene in adult specimens. The northern population of amphipods showed a greater magnitude of induction and a 2 °C lower onset temperature when compared to the southern population, suggesting that the northern population is more affected by elevated temperature than the southern one. We demonstrated that the expression of hsp70 may play a crucial role in the persistence of biogeographical patterns of G. pulex, since it reflects the natural distribution of this species along the latitudinal thermal gradient. A differential regulation of hsc70 gene was also observed according to the ontogenetic stage, with a switch from heat inducible in early life stages to constitutively and highly expressed in adults. These findings demonstrate the importance of considering the entire life cycle to better understand the adaptive response to thermal stress.

Is the deep-sea crab Chaceon affinis able to induce a thermal stress response?

Fluctuations in the stress level of an organism are expressed in behavioural and molecular changes that can affect its ecology and survival. Our knowledge of thermal adaptations in deep-sea organisms is very limited, and this study investigates the critical thermal maximum (CTmax) and the heat-shock response (HSR) in the deep-sea crab Chaceon affinis commonly found in waters of the North East Atlantic. A mild but significant HSR in C. affinis was noted and one of the lowest CTmax known amongst Crustacea was revealed (27.5 °C at 0.1 MPa; 28.5 °C at 10 MPa). The thermal sensitivity of this species appears to be reduced at in situ pressure (10 MPa), given the slightly higher CTmax and the significant 3-fold induction of stress genes hsp70 form 1 and hsp70 form 2. Although C. affinis deep-sea habitat is characterized by overall low temperature this species appears to have retained its ability to induce a HSR. This capability may be linked with C. affinis' occasional exploitation of warmer and thermally instable hydrothermal vent fields, where it has been found foraging for food.

Sustained hydrostatic pressure tolerance of the shallow water shrimp Palaemonetes varians at different temperatures: insights into the colonisation of the deep sea.

We investigated the tolerance of adult specimens of the shallow-water shrimp Palaemonetes varians to sustained high hydrostatic pressure (10 MPa) across its thermal tolerance window (from 5 to 27 °C) using both behavioural (survival and activity) and molecular (hsp70 gene expression) approaches. To our knowledge, this paper reports the longest elevated hydrostatic pressure exposures ever performed on a shallow-water marine organism. Behavioural analysis showed a 100% survival rate of P. varians after 7 days at 10 MPa and 5 or 10 °C, whilst cannibalism was observed at elevated temperature (27 °C), suggesting no impairment of specific dynamic action. A significant interaction of pressure and temperature was observed for both behavioural and molecular responses. Elevated pressure was found to exacerbate the effect of temperature on the behaviour of the animals by reducing activity at low temperature and by increasing activity at high temperature. In contrast, only high pressure combined with low temperature increased the expression of hsp70 genes. We suggest that the impressive tolerance of P. varians to sustained elevated pressure may reflect the physiological capability of an ancestral species to colonise the deep sea. Our results also support the hypothesis that deep-sea colonisation may have occurred during geological periods of time when the oceanic water column was warm and vertically homogenous.

Disentangling the effects of local and regional factors on the thermal tolerance of freshwater crustaceans.

In the global warming context, we compared the thermal tolerance of several populations of the crustacean Gammarus pulex (Amphipoda: Gammaridae) along a latitudinal thermal gradient in the Rhône Valley. To disentangle the effect of regional (North vs. South) and local (site-specific) factors, the ecophysiological responses of populations were investigated at two levels of biological organisation: whole organism level considering body size [critical thermal maximum (CTmax), mean speed of locomotion (MS), time mobile (TM)] and organelle function level [mitochondrial respiratory control ratios (RCRs)]. CTmax and RCRs, but not MS and TM, revealed a significantly higher thermal tolerance in southern populations compared to northern ones. Nevertheless, temperatures ≥ 30°C were deleterious for all populations, suggesting that populations located in the warmer limit of the species distribution will be more threatened by climate change as they live closer to their upper thermal limits. The strong differences observed between populations indicate that the species-level thermal tolerance used in predictive models may not be informative enough to study the impact of global warming on species distributions. This work also reveals that an appropriate choice of indicators is essential to study the consequences of global warming since the response of organisms at the whole body level can be influenced by local conditions.

Identification of differentially expressed genes in the hydrothermal vent shrimp Rimicaris exoculata exposed to heat stress.

The deep-sea vent shrimp Rimicaris exoculata dominates the vagile megafauna at most vent sites along the Mid-Atlantic Ridge. This shrimp swarms around the hot end of the hydrothermal biotope where temperature can exceed its critical maximal temperature (33-38.5 ± 2°C). It may therefore be subjected to a thermal regime that is assumed to be stressful for animals. In this study, we used a global transcriptomic approach by constructing suppression subtractive hybridization cDNA libraries in order to identify specific up- and down-regulated genes in R. exoculata exposed to a severe heat stress (1h at 30°C). A total of 218 sequences representing potentially highly expressed genes in thermally stressed shrimp were obtained. Expression of 11 genes involved in various cell functions was quantified in control and heat shocked specimens using real-time PCR. Differential expression was observed for some specific genes such as mannose receptor C1, metalloprotease, histone H1, and hemocyanin with a strong up-regulation of several genes encoding heat shock proteins. These results suggest that R. exoculata is affected at both cellular and molecular levels by sustained exposure at 30°C. The sequenced ESTs presented here will provide an excellent basis for future thermal stress studies on deep-sea vent fauna.

Thermal biology of the deep-sea vent annelid Paralvinella grasslei: in vivo studies.

The annelid Paralvinella grasslei is a deep-sea vent endemic species that colonizes the wall of active chimneys. We report here the first data on its thermal biology based on in vivo experiments in pressurized aquaria. Our results demonstrate that P. grasslei survives a 30 min exposure at 30 degrees C, and suggest that the upper thermal limit of this species is slightly above this temperature. The first signs of stress were noticed at 30 degrees C, such as a significant increase in the animal's activity and the expression of HSP70 stress proteins. A preliminary investigation of the kinetics of stress protein expression surprisingly showed high levels of HSP70 proteins as late as 3.5 h after the heat shock. Finally, we provide here the first sequences for vent annelid hsp70 (P. grasslei, Hesiolyra bergi and Alvinella pompejana). These constitute valuable tools for future studies on the thermal biology of these annelids.

Temperature resistance studies on the deep-sea vent shrimp Mirocaris fortunata.

The shrimp Mirocaris fortunata is a hydrothermal vent species that is found at most vent-sites along the Mid-Atlantic Ridge. This endemic species is found across a hydrothermal gradient, with thermal conditions ranging from 2-9 degrees C in ambient seawater to fairly warm values of about 25 degrees C. We performed in vivo experiments on M. fortunata specimens originating from different sites and depths (850 m to 2300 m), both at atmospheric pressure and in pressurized aquaria, to characterise the upper thermal limits of this species. Atmospheric pressure results show that thermal physiology should be studied at each population's native pressure. At in situ pressure, shrimps from Menez Gwen (850 m depth) and Lucky Strike (1700 m depth) do not survive temperatures of 39 degrees C, and the 'loss of equilibrium' response suggests that their critical thermal maximum (Ctmax), is about 36+/-1 degrees C for both sites. This value is similar to those found for another vent shrimp, Rimicaris exoculata, which is thought to be a more temperature-resistant organism, so temperature resistance does not appear to be a crucial factor for explaining differences in distribution of shrimp species in a given vent site. Finally, the data for both vent shrimps are also comparable to those of other non-vent tropical caridean species.