Description: The strain of Alexandrium ostenfeldii was isolated from a single cell from Ouwerkerkse Kreek.

Description: Temperature dependent growth is an important indicator to understand the thermal tolerance of organisms and to project their vulnerability to future climate change. Direct measurements of temperature dependent weight gains, however, are experimentally challenging and time consuming in long-lived species. Here, we reassess methodology to quantify the in vivo protein synthesis rate from amino acids, as a key component of growth. We developed an analytical method that is both robust against analytical errors and does not require hazardous radioactive materials. We utilized the incorporation of isotopically 13C15N-labeled-phenylalanine into fish muscle followed by quantification by liquid chromatography mass spectrometry to calculate accurate net protein synthesis rates in muscle tissue of Antarctic fish, Pachycara brachycephalum, in vivo. Specifically, we injected 150 mM of 13C9H915N1 phenylalanine intraperitoneally and sampled muscle tissue in 1,5h steps between 0 and 6 hours after injection. We quantified labeled and unlabeled phenylalanine both in muscle protein and in the cytosol. This allowed us to critically re-evaluate three different protein synthesis rate (Ks) calculation methodologies that have been developed over the last decades. The calculated values differ by more than 70-fold (0.048 ± 0.021% day-1 up to 3.56 ± 2.16 day-1) between methods. We argue that the Ks calculation including a proportionate ratio of protein synthesis from (unlabeled) free amino acids yields the most realistic Ks values for cold water fish. Eventually, the standardization of the net protein synthesis rate calculation will lead to dependable quantitative representations of organismal stress in response to climate change.

Description: We measured the algal storage polysaccharide laminarin and particulate organic carbon (POC) concentrations extracted from particulate organic matter collected during six cruises to the Arctic in 2016 and 2017, North Atlantic, Peru upwelling, Canary upwelling, meridional Atlantic transect, in the Raunefjorden near Bergen, and during two time series in the North Sea near the island Helgoland, resulting in a total of over 250 samples from 51 stations. The majority of samples were taken in surface waters between 0 and 40 m depth. Only in the two Arctic and the North Atlantic datasets, samples were obtained from water depths of 350 m below the surface. In parts of the sample set we additionally determined particulate organic nitrogen, chlorophyll a, protein or lipid concentrations.

Description: Protein turnover is highly energy consuming and overall relates to an organism's growth performance varying largely between species, e.g., due to pre-adaptation to environmental characteristics such as temperature. Here, we determined protein synthesis rates and capacity of protein degradation in white muscle of the cold stenothermal Antarctic eelpout (Pachycara brachycephalum) and its closely related temperate counterpart, the eurythermal common eelpout (Zoarces viviparus). Both species were exposed to acute warming (P. brachycephalum, 0 °C + 2 °C/day; Z. viviparus, 4 °C + 3 °C/day). The in vivo protein synthesis rate (Ks) was monitored after injection of 13C-phenylalanine, and protein degradation capacity was quantified by measuring the activity of cathepsin D in vitro. Untargeted metabolic profiling by nuclear magnetic resonance (NMR) spectroscopy was used to identify the metabolic processes involved. Independent of temperature, the protein synthesis rate was higher in P. brachycephalum (Ks = 0.38–0.614 %/day) than in Z. viviparus (Ks= 0.148-0.379%/day). Whereas protein synthesis remained unaffected by temperature in the Antarctic species, protein synthesis in Z. viviparus increased to near the thermal optimum (16 °C) and tended to fall at higher temperatures. Most strikingly, capacities for protein degradation were about ten times higher in the Antarctic compared to the temperate species. These differences are mirrored in the metabolic profiles, with significantly higher levels of complex and essential amino acids in the free cytosolic pool of the Antarctic congener. Together, the results clearly indicate a highly cold-compensated protein turnover in the Antarctic eelpout compared to its temperate confamilial. Constant versus variable environments are mirrored in rigid versus plastic functional responses of the protein synthesis machinery.

Description: Transcripts of relevant enzymes of the storage lipid metabolism were obtained from already existing transcriptomes that are available in the NCBI database (https://blast.ncbi.nlm.nih.gov/Blast.cgi). Transcriptomes originate from pooled midgut gland samples of several individuals cought in the North Sea. The transcriptome of Crangon crangon is found in the BioProject PRJNA479562 and of Pandalus montagui under BioProject PRJNA798226. Transcripts identified as relevant enzymes are noted by their Trinity_ID and identified through Blastn and Blastp. The nucleotide sequences were translated into amino acid sequences wih the largest ORF. More information about each of the transcriptomes is provided under the individual BioProjects.

Description: The storage lipid metabolism of two caridean shrimp species (Crangon crangon and Pandalus montagui) was studied through a combination of enzyme assays, total lipid determination and transcriptome analyses. The initital sampling was carried out in June, July and August 2021 by the research vessel R/V Uthörn. Freshly caught shrimps from the North Sea were measured, weighted and dissected and brought back to the laboratory facilites of the Alfread-Wegener-Institute, Bremerhaven, Germany. Individual midgut glands were weighted to determine the wet mass and freeze-dried. Dry mass was termined and lipids were extracted after Hagen (2000, see in Postel et al. 2000). The total lipid content of individual midgut glands of Crangon crangon and Pandalus montagui was determined gravimetrically. The synthesis of the storage lipid triacylglycerol (TAG) was measured in pooled microsomal fraction of midgut gland tissue of both shrimp species through the activity of the enzyme diacylglycerol acyltransferase (DGAT) at 37 °C in a water bath (McFie & Stone 2011). Here a fluorescent activated fatty acid (NBD-palmitoyl-CoA, 810229 Avanti Polar Lipids) was used. Lipids in the reaction mix were extracted and lipid classes separated on a thin layer chromatography plate. DGAT activity was measured through arbitrary fluorescent units (AFU/min/mg protein) of the correscponding TAG product. Annotated transcriptomes of both species (C. crangon Bioproject: PRJNA479562, NCBI; P. montagui Bioproject PRJNA798226, NCBI) were screened for enzymes involved in the lipid metabolism. Transcripts identified as relevant enzymes using BLAST were translated into amino acid sequences.