Identification and bioactivity of a granulocyte colony-stimulating factor b homologue from large yellow croaker (Larimichthys crocea)
Identification and bioactivity of a granulocyte colony-stimulating factor b homologue from large yellow croaker (Larimichthys crocea)
复制标题
大黄鱼 (Larimichthys crocea) 粒细胞集落刺激因子 b 同源物的鉴定及其生物活性。
DOI：
10.1016/j.fsi.2019.04.048
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发表时间：
2019-07-01
影响因子：
4.7
通讯作者：
Chen, Xinhua
中科院分区：
文献类型：
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作者：
Li, Qiuhua;Xu, Libing;Chen, Xinhua
粒细胞集落刺激因子（GCSF）是一种多效性细胞因子，在哺乳动物的造血、先天性和适应性免疫反应中起关键作用。然而，GCSF在硬骨鱼中的生物活性在很大程度上仍然未知。本研究利用RACE-PCR技术克隆了大黄鱼GCSFb的一个同源基因（LcGCSFb）。LcGCSFb的开放阅读框（ORF）长603 bp，编码200个氨基酸的前体蛋白，其中信号肽19个氨基酸，成熟肽181个氨基酸。在健康鱼中，LcGCSFb在所有检查的组织中组成型表达，在粘液组织中水平最高，如鳃、肠和胃。溶藻弧菌攻击诱导其在头肾、脾脏、鳃、肠和胃中的转录。LcGCSFb转录物也在原代头肾白细胞（PKL）、原代头肾巨噬细胞（PKM）、原代头肾粒细胞（PKG）和头肾细胞系（LYCK）中检测到，并且被灭活的溶藻弧菌显著上调。这些数据表明LcGCSFb可能在抗细菌感染的免疫应答中起作用。重组LcGCSFb蛋白（rLcGCSFb）的体内给药显著上调了炎性细胞因子（IL-6和TNF α）和转录因子C/EBP β的表达水平，这是嗜中性粒细胞增殖所必需的。此外，rLcGCSFb还能增强PKL的吞噬功能。LcGCSFb可能通过促进炎症反应和白细胞吞噬活性参与抗菌免疫。据我们所知，这是第一个报告的免疫调节作用的GCSF在硬骨鱼。
Granulocyte colony-stimulating factor (GCSF) is a pleiotropic cytokine that plays a key role in regulation of hematopoiesis, innate and adaptive immune responses in mammals. However, bioactivity of GCSF in teleost fish remains largely unknown. In this study, a GCSFb homologue from large yellow croaker (Larimichthys crocea) (LcGCSFb) was cloned by RACE-PCR techniques. The open reading frame (ORF) of LcGCSFb is 603 bp long and encoded a protein precursor of 200 amino acids (aa), with a 19-aa signal peptide and a 181-aa mature peptide. In healthy fish, the LcGCSFb was constitutively expressed in all examined tissues, with the highest levels in mucous tissues, such as gills, intestine, and stomach. Its transcripts in head kidney, spleen, gills, intestine and stomach were significantly induced by Vibrio alginolyticus challenge. LcGCSFb transcripts were also detected in primary head kidney leukocytes (PKL), primary head kidney macrophages (PKM), primary head kidney granulocytes (PKG) and head kidney cell line (LYCK), and markedly upregulated by inactivated V. alginolyticus. These data suggested that LcGCSFb may play a role in immune response against bacterial infection. In vivo administration of recombinant LcGCSFb protein (rLcGCSFb) significantly upregulated the expression levels of the inflammatory cytokines (IL-6 and TNF alpha), and transcription factor C/EBP beta, which is required for proliferation of neutrophils. Furthermore, rLcGCSFb showed an ability to strengthen the phagocytosis of PKL in vitro. Taken together, LcGCSFb may be involved in antibacterial immunity via promoting the inflammatory response and the phagocytic activity of leukocytes. To our knowledge, this is the first report on immunoregulatory roles of GCSF in teleost.