Molecular evolution and functional characterization of SOCS3a and SOCS3b in miiuy croaker (Miichthys miiuy)
Introduction
Cytokines play crucial roles in homeostasis maintenance and immune regulation by binding to specific cell surface receptors, which can set off a cascade of processes and elicit downstream effects from responsive cells (Ilangumaran et al., 2004; Rico-Bautista et al., 2006; Jin et al., 2008). Cytokines, including hematopoietic growth factors, interferons (IFNs), and interleukins, can activate the Janus kinase-signal transducer and activator of transcription (JAK-STAT) signal transduction pathway (Yoshimura et al., 2007). Although the JAK-STAT signal transduction pathway is crucial in immune regulation, excessive signal regulation may cause damage to cellular functions and normal homeostasis (Shuai and Liu, 2003). Therefore, a number of proteins have been introduced to regulate the cytokine signaling and thereby prevent disorders pertaining to the initiation of the body's immune response (Yao et al., 2015).
The suppressors of the cytokine signaling (SOCS) family are one of the most important inhibitors to cytokine receptor signaling (Alexander, 2002). Since the discovery of the first member of the SOCS family, a total of eight SOCS family members have been found in mammals so far, namely CISH, SOCS1-7 (Yoshimura et al., 1995; Shuai and Liu, 2003). However, for fish, there are at least 12 members of the SOCS family, including 8 known mammalian counterparts (SOCS1-7 and CISH), and 4 fish-specific members SOCS3b, SOCS5b, SOCS8 and SOCS9 (Jin et al., 2008). There are some members of the SOCS family in fish species, such as SOCS1, SOCS3, and CISH, which are cloned from different fish species, such as zebrafish (Danio rerio), Tetraodon nigroviridis, rainbow trout (Oncorhynchus mykiss), and grass carp (Ctenopharyngodon idella) (Jin et al., 2007a, b; Wang et al., 2010, 2011; Xiao et al., 2010). As reported, these members of SOCS family share similar structures, which include a central SH2 domain and a conserved C-terminal SOCS box (Endo et al., 1997; Naka et al., 1997). Most SOCS proteins act as an inhibitor of the signal transduction pathway by binding to phosphorylated tyrosine residues through their conservative SH2 domains (Krebs and Hilton, 2001; Wang et al., 2011). The proteins of the SOCS family, such as IL-6 (Starr et al., 1997), LPS and TNFα (Bode et al., 1999), and TGF-β (Fox et al., 2003), can be induced by various stimuli.
Among these members of the SOCS family, SOCS1 and SOCS3 are the most studied molecules and play crucial roles in innate and adaptive immunity (Carow and Rottenberg, 2014; Wong et al., 2006). In mammals, many studies have reported that different from SOCS1 that is a powerful inhibitor of JAK-STAT signaling, the most important function of SOCS3 is the inhibitor of signaling by the IL-6 family (Shuai and Liu, 2003). In fish, SOCS1 has been found to be induced by IFN-γ in fibroid RTG-2 and macrophage RTS-11 cells (Wang et al., 2010). Furthermore, the expression of SOCS1 could be induced by IFN-ϕ1 in zebrafish embryos (Lopez-Munoz et al., 2011). With regarding to SOCS3, increasing evidences have shown that SOCS3 is essential for placental trophoblast differentiation and fetal erythropoiesis (Ilangumaran et al., 2004), and it is also considered to be a major regulator of IL-23-mediated STAT3 phosphorylation and Th17 generation (Chen et al., 2006). SOCS3 has also been shown to inhibit JAK-STAT and other signaling pathways, such as NF-кB and MAPK (Baker et al., 2009) in mammals. In fish, several studies have described the molecular of SOCS3 in different fish species, and some studies have also indicated that the expression of SOCS3 can be modulated by IFN-γ and IL-1β in rainbow trout (Wang and Secombes, 2008). Despite the above research progress, the biological functions of SOCS3 in teleost fish remain poorly understood.
Miiuy croaker (Miichthys miiuy) is a representative species of the Sciaenidae family, which is one of the important marine fish. However, the aquaculture industry of the species has suffered huge losses due to various pathogens infection in recent decades (Xu et al., 2012). Miiuy croaker has currently become an excellent model for studying fish immune responses due to extensively studying of the species from transcriptome and whole genome (Che et al., 2014; Chu et al., 2017a; Xu et al., 2016) to immune genes (Shu et al., 2016; Wang et al., 2016; Chu et al., 2017b; Xu et al., 2018). In the present study, two types of SOCS3, namely mmSOCS3a and mmSOCS3b, were identified from miiuy croaker, and further studies were conducted in terms of their genomic organizations, gene structures, gene synteny, and expression analysis. Additionally, studies on the cellular localization and regulatory functions of mmSOCS3a and mmSOCS3b have indicated that they have negatively regulatory roles in activation of ISRE signaling pathways induced by IFNγ- or IFNα, which can enrich knowledge about the functions of SOCS3 gene in teleost fish. Accordingly, this study can not only provide insights into the SOCS family in fish species, but provides an evidence about the negatively regulation of ISRE signaling pathway in teleost fish.
Section snippets
Samples
Healthy miiuy croakers were obtained from the Zhoushan Fisheries Research Institute (Zhejiang, China), then we temporarily placed them in inflatable seawater tanks at 25 °C. After acclimatizing, only healthy fish will be used in the challenged experiments as we described in the previous studies (Xu et al., 2012). These fish samples were randomly divided into control groups and challenged groups. Eight tissues (liver, spleen, kidney, muscle, gill, intestine, heart and skin) were collected from
Characteristics of mmSOCS3a and SOCS3b genes
The mmSOCS3a and mmSOCS3b genes were identified, and the complete cDNA sequences of the genes were deposited in GenBank under accession no. MH078493 and MH078494, respectively. The obtained full length of the mmSOCS3a gene was 2576 bp, and its complementary DNA (cDNA) is 2010 bp and consisted of a 618 bp coding sequence (CDS), a 312 bp 5′-untranslated region (UTR) and a 1080 bp 3′-UTR (Suppl Fig. 1A). The 3′-UTR sequences of mmSOCS3a included three RNA instability motif “ATTTA” sequences and
Discussion
An increasing number of SOCS family genes have been identified in teleost fish, indicating that SOCS genes are ubiquitous in vertebrates (Nie et al., 2014). However, specific studies on teleost SOCS functions are still limited. The conserved functional domains of fish SOCS genes are homologous with mammalian SOCS gene in its structure (Zhao et al., 2017). In the present study, mmSOCS3a and mmSOCS3b were identified and characterized. By studying the structures and analyzing the phylogeny and
Acknowledgements
This study was supported by the National Key Research and Development Project, China (2018YFD0900503) and the National Natural Science Foundation of China, China (31822057).
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These authors contributed equally.