Cloning and characterization of phospholipase D from olive flounder (Paralichthys olivaceus).
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.
Jeon
Soo Jin
SJ
Department of Aquatic Life Medicine, Pukyong National University, Busan 608-737, Republic of Korea.
Kim
Moo-Sang
MS
Ahn
Sang Jung
SJ
Seo
Jung Soo
JS
Lim
Sang Uk
SU
Sung
Ji Hea
JH
Kim
Na Young
NY
Jeong
Hyun Do
HD
Lee
Hyung Ho
HH
Chung
Joon Ki
JK
D000818:Animals; D002460:Cell Line; D003001:Cloning, Molecular; D005432:Flounder / Q000235:genetics*; D015971:Gene Expression Regulation, Enzymologic; D008969:Molecular Sequence Data; D010739:Phospholipase D / Q000235:genetics*; D010802:Phylogeny; D017386:Sequence Homology, Amino Acid
D010739:Phospholipase D