Post-Translational Protein Deimination Signatures in Plasma and Plasma EVs of Reindeer (Rangifer tarandus)

Abstract

The reindeer (caribou) Rangifer tarandus is a Cervidae in the order Artiodactyla. Reindeer are sedentary and migratory populations with circumpolar distribution in the Arctic, Northern Europe, Siberia and North America. Reindeer are an important wild and domesticated species, and have developed various adaptive strategies to extreme environments. Importantly, deer have also been identified to be putative zoonotic carriers, including for parasites, prions and coronavirus. Therefore, novel insights into immune-related markers are of considerable interest. Peptidylarginine deiminases (PADs) are a phylogenetically conserved enzyme family which causes post-translational protein deimination by converting arginine into citrulline in target proteins. This affects protein function in health and disease. Extracellular vesicles (EVs) participate in cellular communication, in physiological and pathological processes, via transfer of cargo material, and their release is partly regulated by PADs. This study assessed deiminated protein and EV profile signatures in plasma from sixteen healthy wild female reindeer, collected in Iceland during screening for parasites and chronic wasting disease. Reindeer plasma EV profiles showed a poly-dispersed distribution from 30 to 400 nm and were positive for phylogenetically conserved EV-specific markers. Deiminated proteins were isolated from whole plasma and plasma EVs, identified by proteomic analysis and protein interaction networks assessed by KEGG and GO analysis. This revealed a large number of deimination-enriched pathways for immunity and metabolism, with some differences between whole plasma and EVs. While shared KEGG pathways for whole plasma and plasma EVs included complement and coagulation pathways, KEGG pathways specific for EVs were for protein digestion and absorption, platelet activation, amoebiasis, the AGE–RAGE signaling pathway in diabetic complications, ECM receptor interaction, the relaxin signaling pathway and the estrogen signaling pathway. KEGG pathways specific for whole plasma were pertussis, ferroptosis, SLE, thyroid hormone synthesis, phagosome, Staphylococcus aureus infection, vitamin digestion and absorption, and prion disease. Further differences were also found between molecular function and biological processes GO pathways when comparing functional STRING networks for deiminated proteins in EVs, compared with deiminated proteins in whole plasma. This study highlights deiminated proteins and EVs as candidate biomarkers for reindeer health and may provide information on regulation of immune pathways in physiological and pathological processes, including neurodegenerative (prion) disease and zoonosis.

Extracellular Vesicle Signatures and Post-Translational Protein Deimination in Purple Sea Urchin (Strongylocentrotus purpuratus) Coelomic Fluid—Novel Insights into Echinodermata Biology

“…Furthermore, 85 arginines, potentially targets of deimination, are found in the 1699 aa protein sequence. This highlights that C3 is very likely to undergo deimination and C3 deimination was recently reported for the first time in teleost fish [ 23 ]; it has since been identified in a range of taxa including elasmobranchs [ 24 ], reptiles [ 25 ], birds [ 47 ], cetaceans [ 50 ], pinnipeds [ 51 ], and artiodactyls [ 52 , 53 ]. Furthermore, in teleost fish, C3 has been found in deiminated form in both serum as well as in serum EVs [ 44 , 46 ] and in mucosal EVs [ 42 ], while in the current study C3 was a deimination candidate in coelomic fluid only, but not in the EVs, while C3 was identified as part of the total proteomic cargo in the EVs.…”

“…The KEGG pentose phosphate pathway was previously identified for deiminated proteins from cetacean sera [50] and in alveolates [31]. The biosynthesis of amino acids KEGG pathway was previously identified for deiminated proteins from bovine plasma and plasma-EVs [52], plasma and plasma-EVs of reindeer [53], naked mole-rats [48], and lampreys [41], from cetacean sera [50], and lobster haemolymph [39]. The fructose and mannose metabolism KEGG pathway was identified for deiminated proteins in cetacean sera [50], lamprey plasma [41], and in alveolates [31].…”

“…The protein processing in ER pathway was previously also identified for deiminated proteins in lamprey plasma-EVs [ 41 ]. The phagosome KEGG pathway was previously identified for deiminated proteins in reindeer plasma [ 53 ], bovine serum and serum-EVs [ 52 ], cetacean sera [ 50 ], and in lobster haemolymph [ 39 ].…”

“…A recent comparative body of research has focused on identifying putative roles for PADs/ADI and downstream deimination of proteins involved in shaping immunity and metabolism in a wide range of taxa throughout the phylogenetic tree, including Alveolata [ 31 ], Mollusca [ 38 ], Crustacea [ 39 ], Merostomata [ 40 ], Agnatha [ 41 ], Chondrichthyes [ 24 ], Teleosts [ 23 , 42 , 43 , 44 , 45 , 46 ], Reptilia [ 25 ], Aves [ 47 ], Rodents [ 48 , 49 ], and sea and land mammals including Cetaceans [ 50 ], Pinnipeds [ 51 ], Artiodactyla [ 52 , 53 ], and Camelidae [ 54 ]. Through these studies, our group has particularly focused on identifying deimination signatures in circulatory fluid (e.g., plasma, serum, haemolymph or coelomic fluid, depending on species) and on characterising circulating extracellular vesicles (EVs), including with respect to their protein, including deiminated protein (and in some cases microRNA) cargos.…”

Molecular Dynamics Study of Citrullinated Proteins Associated with the Development of Rheumatoid Arthritis

“…The authors do not comment on the role of the identified proteins in the pathogenesis of joint diseases. In addition, Marco Sancandi and Stefania D’Alessio showed the presence of a citrullinated form of the AMBP protein in pre-motor Parkinson’s disease and health assessment in animal models [ 57 , 58 ]. Using Western blotting methods and following-up LC-MS/MS analysis, Hoshimi Kawaguchi et al demonstrated the increased serum level of citrullatedAMBP protein in patients with RA.…”