ATP-induced reversed thermal sensitivity of O2 binding in both major haemoglobin polymorphs of the non-endothermic Atlantic cod, Gadus morhua.

Atlantic cod is a species that is affected by climate change, with some populations being exposed to higher temperatures than others. The two polymorphs of its major haemoglobin type (HbI) show an inverse change in frequency along a latitudinal temperature cline in the North East Atlantic, which has been associated with differences in population performance and behavioural traits. An earlier study at the northern distribution limit of the species reported differential temperature sensitivities of red blood cell oxygen (O2) affinity between the northern cold-water HbI-2 polymorph and its southern, warm-water HbI-1 counter-part, which has since widely been held as adaptive for the species across its distributional range. The present study critically re-examined this hypothesis by comparing the thermal sensitivity of O2 binding in both purified HbI polymorphs from the southern, high-temperature distribution limit of the species under controlled conditions of allosteric modifiers of Hb function. Contrary to the prevailing view, the O2 affinity of the major HbI polymorphs did not differ from each other under any of the tested conditions. Depending on pH and ATP concentration, the temperature-sensitive and temperature-insensitive Hb-O2 affinity phenotypes - previously exclusively ascribed to HbI-1 and HbI-2, respectively - could be induced in both HbI polymorphs. These results are the first to establish a molecular mechanism behind a reversed temperature dependence of red blood cell O2 affinity in a non-endotherm fish and lay the basis for future studies on alternative mechanisms behind the differences in distribution, performance and behavioural traits associated with the different HbI polymorphs of Atlantic cod.

Life on the edge: O2 binding in Atlantic cod red blood cells near their southern distribution limit is not sensitive to temperature or haemoglobin genotype.

Atlantic cod are a commercially important species believed to be threatened by warming seas near their southern, equatorward upper thermal edge of distribution. Limitations to circulatory O2 transport, in particular cardiac output, and the geographic distribution of functionally different haemoglobin (Hb) genotypes have separately been suggested to play a role in setting thermal tolerance in this species. The present study assessed the thermal sensitivity of O2 binding in Atlantic cod red blood cells with different Hb genotypes near their upper thermal distribution limit and modelled its consequences for the arterio-venous O2 saturation difference, Sa-vO2 , another major determinant of circulatory O2 supply rate. The results showed statistically indistinguishable red blood cell O2 binding between the three HbI genotypes in wild-caught Atlantic cod from the Irish Sea (53° N). Red blood cells had an unusually low O2 affinity, with reduced or even reversed thermal sensitivity between pH 7.4 and 7.9, and 5.0 and 20.0°C. This was paired with strongly pH-dependent affinity and cooperativity of red blood cell O2 binding (Bohr and Root effects). Modelling of Sa-vO2  at physiological pH, temperature and O2 partial pressures revealed a substantial capacity for increases in Sa-vO2  to meet rising tissue O2 demands at 5.0 and 12.5°C, but not at 20°C. Furthermore, there was no evidence for an increase of maximal Sa-vO2  with temperature. It is suggested that Atlantic cod at such high temperatures may solely depend on increases in cardiac output and blood O2 capacity, or thermal acclimatisation of metabolic rate, for matching circulatory O2 supply to tissue demand.