Wan Junjie, Zhao Xuelin, Liu Jiqing, Chen Kaiyu, Li Chenghua
State Key Laboratory for Quality and Safety of Agro-products, Ningbo University, Ningbo, 315211, PR China; State-Province Joint Laboratory of Marine Biotechnology and Engineering, Ningbo University, Ningbo, 315211, PR China.
State Key Laboratory for Quality and Safety of Agro-products, Ningbo University, Ningbo, 315211, PR China; State-Province Joint Laboratory of Marine Biotechnology and Engineering, Ningbo University, Ningbo, 315211, PR China.
Fish Shellfish Immunol. 2022 May;124:411-420. doi: 10.1016/j.fsi.2022.04.024. Epub 2022 Apr 21.
Immune cells have many efficient ways to participate in the host immunity, including phagocytosis, which is an important pathway to eliminate pathogens. Only β-integrin-mediated phagocytosis pathways have been confirmed in Apostichopus japonicus. The Src family kinases (SFKs), a class of non-receptor tyrosine kinases plays an important role in the regulation of phagocytic signals in invertebrates. However, the SFK-mediated phagocytic mechanism is largely unknown in A. japonicus. In this study, a novel SFK homologue (AjSrc) with a conservative SH domain, an SH domain, and a tyrosine kinase domain was identified from A. japonicus. Both gene and protein expression of AjSrc and phosphorylation levels increased under Vibrio splendidus challenged, reaching the highest level at 24 h. Knock-down of AjSrc could depress coelomocytes' phagocytosis by 25% compared to the control group. To better understand the mechanism of AjSrc-mediated phagocytosis, focal adhesion kinase (FAK) was identified by a Co-immunoprecipitation experiment to be verified as an interactive protein of AjSrc. The phagocytosis rates of coelomocytes were decreased by 33% and 37% in AjFAK and AjSrc + AjFAK interference groups compared with the control group, respectively. Furthermore, the phosphorylation level of AjFAK was increased and reached the maximum level at 24 h post V. splendidus infection, as the same as that of AjSrc. Our results suggested that AjSrc could mediate V. splendidus-induced coelomocytes' phagocytosis via interacting with AjFAK and co-phosphorylation. This study enriched the mechanism of phagocytosis in echinoderm and provided the new theoretical foundation for disease control of sea cucumber.
免疫细胞有许多参与宿主免疫的有效方式，包括吞噬作用，这是消除病原体的重要途径。在刺参中仅证实了β-整合素介导的吞噬途径。Src家族激酶（SFKs）是一类非受体酪氨酸激酶，在无脊椎动物吞噬信号的调节中起重要作用。然而，SFK介导的吞噬机制在刺参中很大程度上尚不清楚。在本研究中，从刺参中鉴定出一种具有保守SH结构域、SH结构域和酪氨酸激酶结构域的新型SFK同源物（AjSrc）。在灿烂弧菌攻击下，AjSrc的基因和蛋白表达以及磷酸化水平均增加，在24小时达到最高水平。与对照组相比，敲低AjSrc可使体腔细胞的吞噬作用降低25%。为了更好地理解AjSrc介导吞噬作用的机制，通过免疫共沉淀实验鉴定出粘着斑激酶（FAK），并被证实为AjSrc的相互作用蛋白。与对照组相比，AjFAK和AjSrc+AjFAK干扰组体腔细胞的吞噬率分别降低了33%和37%。此外，AjFAK的磷酸化水平增加，并在灿烂弧菌感染后24小时达到最高水平，与AjSrc相同。我们的结果表明，AjSrc可通过与AjFAK相互作用和共同磷酸化介导灿烂弧菌诱导的体腔细胞吞噬作用。本研究丰富了棘皮动物吞噬作用的机制，为海参疾病防治提供了新的理论基础。
