Positive selection in octopus haemocyanin indicates functional links to temperature adaptation.

BACKGROUND: Octopods have successfully colonised the world's oceans from the tropics to the poles. Yet, successful persistence in these habitats has required adaptations of their advanced physiological apparatus to compensate impaired oxygen supply. Their oxygen transporter haemocyanin plays a major role in cold tolerance and accordingly has undergone functional modifications to sustain oxygen release at sub-zero temperatures. However, it remains unknown how molecular properties evolved to explain the observed functional adaptations. We thus aimed to assess whether natural selection affected molecular and structural properties of haemocyanin that explains temperature adaptation in octopods. RESULTS: Analysis of 239 partial sequences of the haemocyanin functional units (FU) f and g of 28 octopod species of polar, temperate, subtropical and tropical origin revealed natural selection was acting primarily on charge properties of surface residues. Polar octopods contained haemocyanins with higher net surface charge due to decreased glutamic acid content and higher numbers of basic amino acids. Within the analysed partial sequences, positive selection was present at site 2545, positioned between the active copper binding centre and the FU g surface. At this site, methionine was the dominant amino acid in polar octopods and leucine was dominant in tropical octopods. Sites directly involved in oxygen binding or quaternary interactions were highly conserved within the analysed sequence. CONCLUSIONS: This study has provided the first insight into molecular and structural mechanisms that have enabled octopods to sustain oxygen supply from polar to tropical conditions. Our findings imply modulation of oxygen binding via charge-charge interaction at the protein surface, which stabilize quaternary interactions among functional units to reduce detrimental effects of high pH on venous oxygen release. Of the observed partial haemocyanin sequence, residue 2545 formed a close link between the FU g surface and the active centre, suggesting a role as allosteric binding site. The prevalence of methionine at this site in polar octopods, implies regulation of oxygen affinity via increased sensitivity to allosteric metal binding. High sequence conservation of sites directly involved in oxygen binding indicates that functional modifications of octopod haemocyanin rather occur via more subtle mechanisms, as observed in this study.

Blue blood on ice: modulated blood oxygen transport facilitates cold compensation and eurythermy in an Antarctic octopod.

INTRODUCTION: The Antarctic Ocean hosts a rich and diverse fauna despite inhospitable temperatures close to freezing, which require specialist adaptations to sustain animal activity and various underlying body functions. While oxygen transport has been suggested to be key in setting thermal tolerance in warmer climates, this constraint is relaxed in Antarctic fishes and crustaceans, due to high levels of dissolved oxygen. Less is known about how other Antarctic ectotherms cope with temperatures near zero, particularly the more active invertebrates like the abundant octopods. A continued reliance on the highly specialised blood oxygen transport system of cephalopods may concur with functional constraints at cold temperatures. We therefore analysed the octopod's central oxygen transport component, the blue blood pigment haemocyanin, to unravel strategies that sustain oxygen supply at cold temperatures. RESULTS: To identify adaptive compensation of blood oxygen transport in octopods from different climatic regions, we compared haemocyanin oxygen binding properties, oxygen carrying capacities as well as haemolymph protein and ion composition between the Antarctic octopod Pareledone charcoti, the South-east Australian Octopus pallidus and the Mediterranean Eledone moschata. In the Antarctic Pareledone charcoti at 0°C, oxygen unloading by haemocyanin was poor but supported by high levels of dissolved oxygen. However, lower oxygen affinity and higher oxygen carrying capacity compared to warm water octopods, still enabled significant contribution of haemocyanin to oxygen transport at 0°C. At warmer temperatures, haemocyanin of Pareledone charcoti releases most of the bound oxygen, supporting oxygen supply at 10°C. In warm water octopods, increasing oxygen affinities reduce the ability to release oxygen from haemocyanin at colder temperatures. Though, unlike Eledone moschata, Octopus pallidus attenuated this increase below 15°C. CONCLUSIONS: Adjustments of haemocyanin physiological function and haemocyanin concentrations but also high dissolved oxygen concentrations support oxygen supply in the Antarctic octopus Pareledone charcoti at near freezing temperatures. Increased oxygen supply by haemocyanin at warmer temperatures supports extended warm tolerance and thus eurythermy of Pareledone charcoti. Limited haemocyanin function towards colder temperatures in Antarctic and warm water octopods highlights the general role of haemocyanin oxygen transport in constraining cold tolerance in octopods.

Simultaneous high-resolution pH and spectrophotometric recordings of oxygen binding in blood microvolumes.

Oxygen equilibrium curves have been widely used to understand oxygen transport in numerous organisms. A major challenge has been to monitor oxygen binding characteristics and concomitant pH changes as they occur in vivo, in limited sample volumes. Here we report a technique allowing highly resolved and simultaneous monitoring of pH and blood pigment saturation in minute blood volumes. We equipped a gas diffusion chamber with a broad-range fibre-optic spectrophotometer and a micro-pH optode and recorded changes of pigment oxygenation along oxygen partial pressure (PO2) and pH gradients to test the setup. Oxygen binding parameters derived from measurements in only 15 µl of haemolymph from the cephalopod Octopus vulgaris showed low instrumental error (0.93%) and good agreement with published data. Broad-range spectra, each resolving 2048 data points, provided detailed insight into the complex absorbance characteristics of diverse blood types. After consideration of photobleaching and intrinsic fluorescence, pH optodes yielded accurate recordings and resolved a sigmoidal shift of 0.03 pH units in response to changing PO2 from 0 to 21 kPa. Highly resolved continuous recordings along pH gradients conformed to stepwise measurements at low rates of pH changes. In this study we showed that a diffusion chamber upgraded with a broad-range spectrophotometer and an optical pH sensor accurately characterizes oxygen binding with minimal sample consumption and manipulation. We conclude that the modified diffusion chamber is highly suitable for experimental biologists who demand high flexibility, detailed insight into oxygen binding as well as experimental and biological accuracy combined in a single setup.

Mitochondrial dynamics underlying thermal plasticity of cuttlefish (Sepia officinalis) hearts.

In the eurythermal cuttlefish Sepia officinalis, performance depends on hearts that ensure systemic oxygen supply over a broad range of temperatures. We therefore aimed to identify adjustments in energetic cardiac capacity and underlying mitochondrial function supporting thermal acclimation and adaptation that could be crucial for the cuttlefish's competitive success in variable environments. Two genetically distinct cuttlefish populations were acclimated to 11, 16 and 21°C. Subsequently, skinned and permeabilised heart fibres were used to assess mitochondrial functioning by means of high-resolution respirometry and a substrate-inhibitor protocol, followed by measurements of cardiac citrate synthase and cytosolic enzyme activities. Temperate English Channel cuttlefish had lower mitochondrial capacities but larger hearts than subtropical Adriatic cuttlefish. Warm acclimation to 21°C decreased mitochondrial complex I activity in Adriatic cuttlefish and increased complex IV activity in English Channel cuttlefish. However, compensation of mitochondrial capacities did not occur during cold acclimation to 11°C. In systemic hearts, the thermal sensitivity of mitochondrial substrate oxidation was high for proline and pyruvate but low for succinate. Oxygen efficiency of catabolism rose as temperature changed from 11 to 21°C via shifts to oxygen-conserving oxidation of proline and pyruvate and via reduced relative proton leak. The changes observed for substrate oxidation, mitochondrial complexes, relative proton leak and heart mass improve energetic efficiency and essentially seem to extend tolerance to high temperatures and reduce associated tissue hypoxia. We conclude that cuttlefish sustain cardiac performance and, thus, systemic oxygen delivery over short- and long-term changes of temperature and environmental conditions by multiple adjustments in cellular and mitochondrial energetics.

Metabolic plasticity improves lobster's resilience to ocean warming but not to climate-driven novel species interactions.

Marine species not only suffer from direct effects of warming oceans but also indirectly via the emergence of novel species interactions. While metabolic adjustments can be crucial to improve resilience to warming, it is largely unknown if this improves performance relative to novel competitors. We aimed to identify if spiny lobsters-inhabiting a global warming and species re-distribution hotspot-align their metabolic performance to improve resilience to both warming and novel species interactions. We measured metabolic and escape capacity of two Australian spiny lobsters, resident Jasus edwardsii and the range-shifting Sagmariasus verreauxi, acclimated to current average-(14.0 °C), current summer-(17.5 °C) and projected future summer-(21.5 °C) habitat temperatures. We found that both species decreased their standard metabolic rate with increased acclimation temperature, while sustaining their scope for aerobic metabolism. However, the resident lobster showed reduced anaerobic escape performance at warmer temperatures and failed to match the metabolic capacity of the range-shifting lobster. We conclude that although resident spiny lobsters optimise metabolism in response to seasonal and future temperature changes, they may be unable to physiologically outperform their range-shifting competitors. This highlights the critical importance of exploring direct as well as indirect effects of temperature changes to understand climate change impacts.

Open Hardware in Science: The Benefits of Open Electronics.

Openly shared low-cost electronic hardware applications, known as open electronics, have sparked a new open-source movement, with much untapped potential to advance scientific research. Initially designed to appeal to electronic hobbyists, open electronics have formed a global "maker" community and are increasingly used in science and industry. In this perspective article, we review the current costs and benefits of open electronics for use in scientific research ranging from the experimental to the theoretical sciences. We discuss how user-made electronic applications can help (I) individual researchers, by increasing the customization, efficiency, and scalability of experiments, while improving data quantity and quality; (II) scientific institutions, by improving access to customizable high-end technologies, sustainability, visibility, and interdisciplinary collaboration potential; and (III) the scientific community, by improving transparency and reproducibility, helping decouple research capacity from funding, increasing innovation, and improving collaboration potential among researchers and the public. We further discuss how current barriers like poor awareness, knowledge access, and time investments can be resolved by increased documentation and collaboration, and provide guidelines for academics to enter this emerging field. We highlight that open electronics are a promising and powerful tool to help scientific research to become more innovative and reproducible and offer a key practical solution to improve democratic access to science.

Connecting to the oceans: supporting ocean literacy and public engagement.

Improved public understanding of the ocean and the importance of sustainable ocean use, or ocean literacy, is essential for achieving global commitments to sustainable development by 2030 and beyond. However, growing human populations (particularly in mega-cities), urbanisation and socio-economic disparity threaten opportunities for people to engage and connect directly with ocean environments. Thus, a major challenge in engaging the whole of society in achieving ocean sustainability by 2030 is to develop strategies to improve societal connections to the ocean. The concept of ocean literacy reflects public understanding of the ocean, but is also an indication of connections to, and attitudes and behaviours towards, the ocean. Improving and progressing global ocean literacy has potential to catalyse the behaviour changes necessary for achieving a sustainable future. As part of the Future Seas project (https://futureseas2030.org/), this paper aims to synthesise knowledge and perspectives on ocean literacy from a range of disciplines, including but not exclusive to marine biology, socio-ecology, philosophy, technology, psychology, oceanography and human health. Using examples from the literature, we outline the potential for positive change towards a sustainable future based on knowledge that already exists. We focus on four drivers that can influence and improve ocean literacy and societal connections to the ocean: (1) education, (2) cultural connections, (3) technological developments, and (4) knowledge exchange and science-policy interconnections. We explore how each driver plays a role in improving perceptions of the ocean to engender more widespread societal support for effective ocean management and conservation. In doing so, we develop an ocean literacy toolkit, a practical resource for enhancing ocean connections across a broad range of contexts worldwide.

Poleward bound: adapting to climate-driven species redistribution.

One of the most pronounced effects of climate change on the world's oceans is the (generally) poleward movement of species and fishery stocks in response to increasing water temperatures. In some regions, such redistributions are already causing dramatic shifts in marine socioecological systems, profoundly altering ecosystem structure and function, challenging domestic and international fisheries, and impacting on human communities. Such effects are expected to become increasingly widespread as waters continue to warm and species ranges continue to shift. Actions taken over the coming decade (2021-2030) can help us adapt to species redistributions and minimise negative impacts on ecosystems and human communities, achieving a more sustainable future in the face of ecosystem change. We describe key drivers related to climate-driven species redistributions that are likely to have a high impact and influence on whether a sustainable future is achievable by 2030. We posit two different futures-a 'business as usual' future and a technically achievable and more sustainable future, aligned with the Sustainable Development Goals. We then identify concrete actions that provide a pathway towards the more sustainable 2030 and that acknowledge and include Indigenous perspectives. Achieving this sustainable future will depend on improved monitoring and detection, and on adaptive, cooperative management to proactively respond to the challenge of species redistribution. We synthesise examples of such actions as the basis of a strategic approach to tackle this global-scale challenge for the benefit of humanity and ecosystems. Supplementary Information: The online version contains supplementary material available at 10.1007/s11160-021-09641-3.

Thermal sensitivity links to cellular cardiac decline in three spiny lobsters.

Understanding mechanisms of thermal sensitivity is key to predict responses of marine organisms to changing temperatures. Sustaining heart function is critical for complex organisms to oxygenate tissues, particularly under temperature stress. Yet, specific mechanisms that define thermal sensitivity of cardiac function remain unclear. Here we investigated whole animal metabolism, cardiac performance and mitochondrial function in response to elevated temperatures for temperate, subtropical and tropical spiny lobster species. While oxygen demands increased with rising temperatures, heart function became limited or declined in all three species of lobsters. The decline in cardiac performance coincided with decreases in mitochondrial efficiency through increasing mitochondrial proton leakage, which predicts impaired compensation of ATP production. Species differences were marked by shifts in mitochondrial function, with the least thermal scope apparent for tropical lobsters. We conclude that acute temperature stress of spiny lobsters, irrespective of their climatic origin, is marked by declining cellular energetic function of the heart, contributing to an increasing loss of whole animal performance. Better understanding of physiological thermal stress cascades will help to improve forecasts of how changing environmental temperatures affect the fitness of these ecologically and commercially important species.

Mismatch of thermal optima between performance measures, life stages and species of spiny lobster.

In an ocean warming hotspot off south-east Australia, many species have expanded their ranges polewards, including the eastern rock lobster, Sagmariasus verreauxi. This species is likely extending its range via larval advection into Tasmanian coastal waters, which are occupied by the more commercially important southern rock lobster, Jasus edwardsii. Here, thermal tolerances of these lobster species at two life stages were investigated to assess how they may respond to warming ocean temperatures. We found that the pattern, optimum and magnitude of thermal responses differed between performance measures, life stages and species. Sagmariasus verreauxi had a warmer optimal temperature for aerobic scope and escape speed than J. edwardsii. However, J. edwardsii had a higher magnitude of escape speed, indicating higher capacity for escape performance. There were also differences between life stages within species, with the larval stage having higher variation in optimal temperatures between measures than juveniles. This inconsistency in performance optima and magnitude indicates that single performance measures at single life stages are unlikely to accurately predict whole animal performance in terms of life-time survival and fitness. However, combined results of this study suggest that with continued ocean warming, S. verreauxi is likely to continue to extend its distribution polewards and increase in abundance in Tasmania.

A new haemocyanin in cuttlefish (Sepia officinalis) eggs: sequence analysis and relevance during ontogeny.

BACKGROUND: Haemocyanin is the respiratory protein of most of the Mollusca. In cephalopods and gastropods at least two distinct isoforms are differentially expressed. However, their physiological purpose is unknown. For the common cuttlefish Sepia officinalis, three isoforms are known so far, whereas for only two of them the complete mRNA sequences are available. In this study, we sequenced the complete mRNA of the third haemocyanin isoform and measured the relative expression of all three isoforms during embryogenesis to reveal a potential ontogenetic relevance. RESULTS: The cDNA of isoform 3 clearly correlates to the known Sepia officinalis haemocyanin subunits consisting of eight functional units and an internal duplicated functional unit d. Our molecular phylogenetic analyses reveal the third isoform representing a potentially ancestral haemocyanin isoform, and the analyses of the expression of haemocyanin type 3 reveal that haemocyanin type 3 only can be observed within eggs and during early development. Isoforms 1 and 2 are absent at these stages. After hatching, isoform 3 is downregulated, and isoform 1 and 2 are upregulated. CONCLUSIONS: Our study clearly shows an embryonic relevance of the third isoform, which will be further discussed in the light of the changes in the physiological function of haemocyanin during ontogeny. Taken together with the fact that it could also be the isoform closest related to the common ancestor of cuttlefish haemocyanin, the phylogeny of cuttlefish haemocyanin may be recapitulated during its ontogeny.