Granulocyte colony-stimulating factor (GCSF) is a growth factor that drives the proliferation and differentiation of granulocytes and monocytes/macrophages. Currently, two copies of GCSF, named GCSFa and GCSFb, have been identified in teleost fish, but data on the functions and signal pathways of these fish GCSFs are still limited. In the present study, a GCSFa homologue (LcGCSFa) was identified from large yellow croaker (Larimichthys crocea). The open reading frame (ORF) of LcGCSFa is 636 bp long and encodes a protein of 211 amino acids (aa), with a 19-aa signal peptide and a typical IL-6 domain, conserved in fish GCSF sequences. The phylogenetic analysis showed that LcGCSFa clustered with other fish GCSFa homologues. LcGCSFa was constitutively expressed in all tissues tested and significantly up-regulated in head kidney and spleen by Vibrio alginolyticus or poly(I:C). LcGCSFa transcripts were also detected in primary head kidney leucocytes (PKL), primary head kidney macrophages (PKM), and primary head kidney granulocytes (PKG), and significantly up-regulated in PKL and PKG by LPS or poly(I:C). These data indicated that LcGCSFa may be involved in the immune responses induced by bacterium and virus. The recombinant LcGCSFa protein (rLcGCSFa) produced in Pichia pastoris promoted the proliferation of PKL both in vivo and in vitro. Furthermore, rLcGCSFa significantly increased both transcription and phosphorylation levels of the signal transducers and activators of transcription (STAT) proteins (LcSTAT3 and LcSTAT5) in PKL, which are required for the GCSF-dependent proliferation. These results showed that LcGCSFa may promote the proliferation of PKL via the activation of LcSTAT3 and LcSTAT5, suggesting a conserved role across vertebrate GCSFs.

中文翻译：

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粒细胞集落刺激因子a从大黄鱼（Larimichthys crocea）的同系物的鉴定和生物活性。

粒细胞集落刺激因子（GCSF）是一种驱动粒细胞和单核细胞/巨噬细胞增殖和分化的生长因子。目前，已经在硬骨鱼中鉴定出两个副本的GCSF，分别称为GCSFa和GCSFb，但是这些鱼类GCSF的功能和信号途径的数据仍然有限。在本研究中，从大型黄花鱼（Larimichthys crocea）中鉴定出GCSFa同源物（LcGCSFa）。LcGCSFa的开放阅读框（ORF）长636 bp，编码211个氨基酸（aa）的蛋白质，具有19-aa信号肽和典型的IL-6结构域，在鱼类GCSF序列中保守。系统发育分析表明，LcGCSFa与其他鱼类GCSFa同源物成簇。LcGCSFa在所有测试的组织中组成性表达，并通过溶藻弧菌或聚（I：C）在头肾和脾脏中显着上调。在初级头肾白细胞（PKL），初级头肾巨噬细胞（PKM）和初级头肾粒细胞（PKG）中也检测到LcGCSFa转录物，并且LPS或poly（I：C）在PKL和PKG中显着上调了LcGCSFa转录本。这些数据表明LcGCSFa可能参与细菌和病毒诱导的免疫反应。在巴斯德毕赤酵母中产生的重组LcGCSFa蛋白（rLcGCSFa）在体内和体外均促进了PKL的增殖。此外，rLcGCSFa显着增加了PKL中信号转导子和转录激活因子（STAT）蛋白（LcSTAT3和LcSTAT5）的转录和磷酸化水平，这是GCSF依赖性增殖所必需的。这些结果表明，LcGCSFa可能通过LcSTAT3和LcSTAT5的激活来促进PKL的增殖，表明在脊椎动物GCSF中的保守作用。