Shenzhen MSU-BIT University, No. 1, International University Park Road, Dayun New Town, Longgang District, Shenzhen 518172, China.
Shemyakin-Ovchinnikov Institute of Bioorganic Chemistry, Russian Academy of Sciences, Miklukho-Maklaya Str. 16/10, 119997 Moscow, Russia.
Mar Drugs. 2024 Oct 30;22(11):488. doi: 10.3390/md22110488.
Three-finger proteins (TFPs), or Ly6/uPAR proteins, are characterized by the beta-structural LU domain containing three protruding "fingers" and stabilized by four conserved disulfide bonds. TFPs were initially characterized as snake alpha-neurotoxins, but later many studies showed their regulatory roles in different organisms. Despite a known expression of TFPs in vertebrates, they are poorly studied in other taxa. The presence of TFPs in starfish was previously shown, but their targets and functional role still remain unknown. Here, we analyzed expression, target, and possible function of the Lystar5 protein from the starfish using bioinformatics, qPCR, and immunoassay. First, the presence of Lystar5 homologues in all classes of echinoderms was demonstrated. qPCR revealed that mRNA of Lystar5 and LyAr2 are expressed mainly in coelomocytes and coelomic epithelium of , while mRNA of other TFPs, LyAr3, LyAr4, and LyAr5, were also found in a starfish body wall. Using anti-Lystar5 serum from mice immunized by a recombinant Lystar5, we confirmed that this protein is expressed on the surface of coelomocytes and coelomic epithelium cells. According to ELISA, a recombinant analogue of Lystar5 bound to the membrane fraction of coelomocytes and coelomic epithelium but not to the body wall or starfish arm tip. Analysis by LC-MALDI MS/MS suggested integrin α-8-like protein expressed in the coelomocytes and coelomic epithelium as a target of Lystar5. Thus, our insights propose the important role of TFPs in regulation of starfish physiology and show prospects for their further research.
三指蛋白（TFPs）或 Ly6/uPAR 蛋白的特征是含有三个突出“手指”的β结构 LU 结构域，并由四个保守的二硫键稳定。TFPs 最初被表征为蛇α-神经毒素，但后来许多研究表明它们在不同生物体中具有调节作用。尽管已知 TFPs 在脊椎动物中有表达，但在其他分类群中研究甚少。以前已经证明海星中存在 TFPs，但它们的靶标和功能作用仍然未知。在这里，我们使用生物信息学、qPCR 和免疫测定法分析了海星的 Lystar5 蛋白的表达、靶标和可能的功能。首先，证明了 Lystar5 同源物存在于所有棘皮动物门的类群中。qPCR 显示，Lystar5 和 LyAr2 的 mRNA 主要在体腔细胞和体腔上皮中表达，而其他 TFPs 的 mRNA，LyAr3、LyAr4 和 LyAr5，也存在于海星体壁中。使用从小鼠免疫重组 Lystar5 获得的抗 Lystar5 血清，我们证实该蛋白在体腔细胞和体腔上皮细胞的表面表达。根据 ELISA，重组 Lystar5 类似物与体腔细胞和体腔上皮的膜部分结合，但不与体壁或海星腕部结合。LC-MALDI MS/MS 分析表明，整合素α-8 样蛋白作为 Lystar5 的靶标在体腔细胞和体腔上皮中表达。因此，我们的研究结果提出了 TFPs 在调节海星生理学中的重要作用，并展示了它们进一步研究的前景。
