Niimi Satoko, Kim Eun-Young, Iwata Hisato, Watanabe Michio X, Yasunaga Genta, Fujise Yoshihiro, Tanabe Shinsuke
Center for Marine Environmental Studies (CMES), Ehime University, Bunkyo-cho 2-5, Matsuyama 790-8577, Japan.
Comp Biochem Physiol B Biochem Mol Biol. 2007 Aug;147(4):667-81. doi: 10.1016/j.cbpb.2007.04.005. Epub 2007 Apr 14.
Full-length cDNA sequences of cytochrome P450 (CYP) 2C78, 2E1, 3A72, 4A35 and 4V6 isozymes were isolated from a hepatic cDNA library of common minke whale (Balaenoptera acutorostrata). The deduced amino acid sequences of minke whale CYP2C78, 2E1, 3A72, 4A35 and 4V6 showed high identities with cattle CYP2C86 (83%), pig CYP2E1 (85%), sheep CYP3A24 (82%), pig CYP4A21 (80%), and human CYP4V2 (76%), respectively. To investigate whether or not these CYP expression levels are altered by contamination of organochlorine contaminants (OCs), mRNA levels of these CYPs in the liver of common minke whale were measured using a quantitative real-time RT-PCR method, and the quantified mRNA levels were employed for the statistical analysis with the residue levels of OCs including PCBs, DDTs (p,p'-DDT, p,p'-DDD and p,p'-DDE), chlordanes (cis-chlordane, trans-chlordane, cis-nonachlor, trans-nonachlor and oxychlordane), HCHs (alpha-, beta- and gamma-isomers) and hexachlorobenzene that have already been reported elsewhere. Spearman's rank correlation analyses showed no significant correlation between CYP expression levels and each OC level in the common minke whale liver, implying that these environmental chemicals have no potential to alter the expression levels of these CYPs or the residue levels encountered in the whale livers may not reach their transcriptional regulation levels. This suggests that the expression of individual CYPs in the whale liver may be at basal level. Relationships among hepatic mRNA expression levels of these CYP2-4 isozymes together with CYP1A1 and CYP1A2 were also examined. Significant positive correlations were detected among mRNA expression levels of individual CYP isozymes in most cases. These associations indicate that the transcriptional regulation of these CYPs examined in this study may be reciprocally related. CYP1A1 levels showed a positive correlation with CYP1A2 levels (r=0.64, p<0.01) indicating that both CYP isozymes were regulated by aryl hydrocarbon receptor activated by endogenous ligands. A strong positive correlation between CYP2C78 and 3A72 (r=0.90, p<0.001) suggests that expression of these CYP isozymes may be under a regulation mechanism of cross-talk in which specific nuclear receptors such as constitutive androstane receptor and pregnane X receptor are involved. The present study indicates that minke whale from the North Pacific may be a model species to investigate the mechanism of basal regulation of these CYPs.
从小须鲸（Balaenoptera acutorostrata）的肝脏cDNA文库中分离出细胞色素P450（CYP）2C78、2E1、3A72、4A35和4V6同工酶的全长cDNA序列。小须鲸CYP2C78、2E1、3A72、4A35和4V6推导的氨基酸序列分别与牛CYP2C86（83%）、猪CYP2E1（85%）、绵羊CYP3A24（82%）、猪CYP4A21（80%）和人CYP4V2（76%）具有高度同源性。为了研究这些CYP的表达水平是否会因有机氯污染物（OCs）的污染而改变，采用定量实时RT-PCR方法测定了小须鲸肝脏中这些CYPs的mRNA水平，并将定量的mRNA水平与包括多氯联苯、滴滴涕（p,p'-滴滴涕、p,p'-滴滴滴和p,p'-滴滴伊）、氯丹（顺式氯丹、反式氯丹、顺式九氯、反式九氯和氧氯丹）、六氯环己烷（α-、β-和γ-异构体）和六氯苯在内的OCs残留水平进行统计分析，这些OCs的残留水平已在其他地方报道过。Spearman等级相关分析表明，小须鲸肝脏中CYP表达水平与每种OC水平之间无显著相关性，这意味着这些环境化学物质没有改变这些CYPs表达水平的潜力，或者鲸鱼肝脏中遇到的残留水平可能未达到其转录调控水平。这表明鲸鱼肝脏中单个CYPs的表达可能处于基础水平。还研究了这些CYP2-4同工酶以及CYP1A1和CYP1A2的肝脏mRNA表达水平之间的关系。在大多数情况下，单个CYP同工酶的mRNA表达水平之间检测到显著的正相关。这些关联表明，本研究中检测的这些CYPs的转录调控可能是相互关联的。CYP1A1水平与CYP1A2水平呈正相关（r = 0.64，p < 0.01），表明这两种CYP同工酶均受内源性配体激活的芳烃受体调控。CYP2C78与3A72之间存在强正相关（r = 0.90，p < 0.001），表明这些CYP同工酶的表达可能受串扰调节机制的调控，其中涉及组成型雄甾烷受体和孕烷X受体等特定核受体。本研究表明，北太平洋的小须鲸可能是研究这些CYPs基础调控机制的模式物种。
