Cloning, expression and functional analysis of a novel-chemokine gene of Japanese flounder, Paralichthys olivaceus, containing two additional cysteines and an extra fourth exon.
Cloning, expression and functional analysis of a novel-chemokine gene of Japanese flounder, Paralichthys olivaceus, containing two additional cysteines and an extra fourth exon.
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DOI：
10.1016/j.fsi.2006.08.012
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发表时间：
2007-06
影响因子：
4.7
通讯作者：
Rutch Khattiya;H. Kondo;I. Hirono;T. Aoki
中科院分区：
文献类型：
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作者：
Rutch Khattiya;H. Kondo;I. Hirono;T. Aoki
从牙鲆（paricichthys olivaceus）中克隆了一个CC趋化因子基因（JFCCL3）并进行了测序。JFCCL3 cDNA包含一个开放阅读框，288个核苷酸编码95个氨基酸残基。预测的JFCCL3氨基酸序列显示β趋化因子的保守半胱氨酸加上两个额外的半胱氨酸。基因组序列由两个亚型组成：JFCCL3.1和JFCCL3.2，大小分别为1.8 kb和1.2kb。这两种异构体都含有三个内含子和四个外显子。RT-PCR结果显示JFCCL3在包括淋巴器官在内的大多数组织中均有组成性表达。通过实时定量RT-PCR，观察到JFCCL3转录本在Con A/PMA刺激后3小时和LPS刺激后1小时在pbl中的最高表达。系统发育分析表明，JFCCL3与fractalkines的亲缘关系比与其他哺乳动物β趋化因子的亲缘关系更密切。趋化性实验表明，重组JFCCL3蛋白在0.01 ~ 10μg/ml浓度范围内具有吸引牙鲆白细胞的生物活性。
A CC chemokine gene (JFCCL3) was cloned and sequenced from Japanese flounder, Paralichthys olivaceus.The JFCCL3 cDNA contains an open reading frame of 288 nucleotides encoding 95 amino acid residues. The predicted amino acid sequence of JFCCL3 showed the conserved cysteine of the β chemokine plus two additional cysteines. The genomic sequence consists of two isoforms: JFCCL3.1 and JFCCL3.2 with sizes of 1.8 and 1.2kb, respectively. Both isoforms contain three introns and four exons. RT–PCR showed that JFCCL3 is constitutively expressed in most tissues including lymphoid organs. Using quantitative real-time RT–PCR, the highest expression of JFCCL3 transcripts was observed in PBLs at 3h post-stimulation with Con A/PMA and at 1h post-stimulation with LPS. A phylogenetic analysis showed that JFCCL3 is more closely related to fractalkines than to other mammalian β chemokines. A chemotaxis assay showed that recombinant JFCCL3 protein has bioactivity for Japanese flounder leukocyte attraction at concentrations from 0.01 to 10μg/ml.