Jeon Soo Jin, Kim Moo-Sang, Ahn Sang Jung, Seo Jung Soo, Lim Sang Uk, Sung Ji Hea, Kim Na Young, Jeong Hyun Do, Lee Hyung Ho, Chung Joon Ki
Department of Aquatic Life Medicine, Pukyong National University, Busan 608-737, Republic of Korea.
Fish Shellfish Immunol. 2008 May;24(5):542-50. doi: 10.1016/j.fsi.2007.11.004. Epub 2007 Nov 21.
The phospholipase D1 (PLD1) cDNA, designated PoPLD, encoding a predicted protein of 1053 amino acids in olive flounder (Paralichthys olivaceus) has been cloned. The deduced amino acid sequence shares high identity with that of PLD1s and PLD2 in human, rat and mouse. The phylogenic analysis and sequence comparison of PoPLD with other PLD isozymes were found to be closely related to the PLD1 isozyme in primary structure. The tissue expression analysis of PoPLD showed that the mRNA of PoPLD was predominantly expressed in the brain, gullet, muscle, stomach, head kidney, pyloric caeca, intestine and gill. The expression of the PoPLD gene was examined in various tissues of flounder by RT-PCR following stimulation with LPS and compared also with that of the inflammatory cytokines IL-1beta and IL-8 in various tissues of the stimulated flounder. This provides indirect evidence that PLD1 might have a relevant role in immune responses against pathogens and in inflammation. In addition, the recombinant protein of PoPLD (GFP-PoPLD), which demonstrated a phosphatidylcholine (PC)-hydrolyzing activity, was partially localized as a distinct ring-shaped form surrounding the rim of the nucleus in EPC cells. Together, our results suggest that PoPLD is similar to the mammalian PLD1 isoform, is generally widespread within olive flounder tissue, might have a relevant role in the fish immune system against pathogens and specifically may be localized in the subcellular membranes of the nuclear rim in EPC cells.
已克隆出牙鲆（Paralichthys olivaceus）中编码预测的1053个氨基酸的蛋白质的磷脂酶D1（PLD1）cDNA，命名为PoPLD。推导的氨基酸序列与人类、大鼠和小鼠的PLD1及PLD2具有高度同源性。发现PoPLD与其他PLD同工酶的系统发育分析和序列比较在一级结构上与PLD1同工酶密切相关。PoPLD的组织表达分析表明，PoPLD的mRNA主要在脑、食道、肌肉、胃、头肾、幽门盲囊、肠道和鳃中表达。在用LPS刺激后，通过RT-PCR检测了牙鲆各组织中PoPLD基因的表达，并将其与受刺激牙鲆各组织中炎性细胞因子IL-1β和IL-8的表达进行了比较。这提供了间接证据，表明PLD1可能在针对病原体的免疫反应和炎症中发挥相关作用。此外，具有磷脂酰胆碱（PC）水解活性的PoPLD重组蛋白（GFP-PoPLD）在EPC细胞中部分定位于围绕细胞核边缘的独特环形结构。总之，我们的结果表明，PoPLD与哺乳动物PLD1亚型相似，在牙鲆组织中普遍存在，可能在鱼类针对病原体的免疫系统中发挥相关作用，并且在EPC细胞中可能特异性定位于核边缘的亚细胞膜。
