Iron has a fundamental role in life and in its biochemical reactions but, when in excess, it can promote the formation of free radicals which can lead to cell death. Therefore, managing the levels of iron is essential to regulate the production of oxidative stress related to iron, and ferritins are one of the main protein families involved in this process. Ferritins are ≈480 kDa multimeric proteins composed by 24 subunits, each with 19−26 kDa, which can accumulate up to 4500 iron atoms. Besides their role in managing iron bioavailability, they have also developed a role in organism immunity and defence present throughout evolution. In this work, we identified and characterized, for the first time, four different ferritin subunits in the clam Ruditapes decussatus, a bivalve commercially and ecologically important along the south Atlantic coast and in the Mediterranean basin, which is a major target of the parasitic protozoa Perkinsus olseni, considered one of the main causes of high levels of clam mortality. Following phylogenetic annotation, the four ferritins subunits identified were subdivided into two cytosolic and two secreted forms. All four subunits maintain the canonical ferritin structure with four main helices α (A–D) and a small helix (E), but the secreted ferritins present an additional helix in their N-terminal region (F), located after the signal peptide and with possible antimicrobial properties. Additionally, we identified in ferritin 4 an extra helix α (G) located between helices B and C. These alpha helix domains revealed high degree of similarity with antimicrobial peptides associated with antibacterial and antifungal activities. Analysis of the expression of these subunits showed that ferritins 1 and 2 are ubiquitously expressed while ferritins 3 and 4 are present mainly in visceral mass. Ferritin 1 lacked a putative functional iron response element (IRE) and appeared to be under a tight regulation. Ferritins 2 and 3 showed a strong response to infection by parasite Perkinsus olseni in contrast to ferritin 4, whose main response was related to exposure to a combination of metals. The synergistic effect between metals and infection promoted a general upregulation of the four ferritins. In conclusion, our results suggest that ferritins, besides their function in iron and metals detoxification, may play a determinant role in clam immune response.

铁在生命及其生化反应中起着基本作用，但是当铁过量时，它会促进自由基的形成，从而导致细胞死亡。因此，控制铁水平对于调节与铁有关的氧化应激的产生至关重要，铁蛋白是该过程中涉及的主要蛋白质家族之一。铁蛋白是约480 kDa的多聚体蛋白，由24个亚基组成，每个亚基具有19-26 kDa，可累积多达4500个铁原子。除了在管理铁生物利用度方面的作用外，他们还在整个进化过程中对机体免疫力和防御力发挥了作用。在这项工作中，我们首次确定了蛤Ru Ruditapes decussatus中的四个不同铁蛋白亚基，并对其进行了表征，沿南大西洋海岸和地中海盆地具有重要商业和生态意义的双壳类动物，这是寄生原生动物Perkinsus olseni的主要目标，被认为是蛤类死亡率高的主要原因之一。在系统发生注释之后，将鉴定出的四个铁蛋白亚基细分为两种胞质和两种分泌形式。所有四个亚基均维持具有四个主要螺旋α（A–D）和一个小螺旋（E）的典型铁蛋白结构，但分泌的铁蛋白在其N末端区域（F）处又有一个螺旋，位于信号肽和具有可能的抗菌特性。此外，我们在铁蛋白4中鉴定出位于螺旋B和C之间的额外螺旋α（G）。这些α螺旋结构域显示与与抗菌和抗真菌活性相关的抗菌肽高度相似。对这些亚基表达的分析表明，铁蛋白1和2普遍存在，而铁蛋白3和4主要存在于内脏肿块中。铁蛋白1缺乏假定的功能性铁反应元件（IRE），并且似乎处于严格的调节之下。铁蛋白2和3对寄生虫感染表现出强烈的反应Perkinsus olseni与铁蛋白4相反，铁蛋白4的主要反应与暴露于金属组合有关。金属与感染之间的协同作用促进了四种铁蛋白的普遍上调。总之，我们的结果表明，铁蛋白除具有铁和金属排毒功能外，还可能在蛤类免疫反应中起决定性作用。