Key Laboratory of Freshwater Animal Breeding Ministry of Agriculture of P.R.China, Fishery College, Huazhong Agricultural University, Wuhan 430070, China.
Key Laboratory of Freshwater Animal Breeding Ministry of Agriculture of P.R.China, Fishery College, Huazhong Agricultural University, Wuhan 430070, China; Collaborative Innovation Center for Efficient and Health Production of Fisheries in Hunan Province, Changde 415000, China; Hubei Provincial Engineering Laboratory for Pond Aquaculture, Fishery College, Huazhong Agricultural University, Wuhan 430070, China.
J Trace Elem Med Biol. 2017 Dec;44:256-265. doi: 10.1016/j.jtemb.2017.09.004. Epub 2017 Sep 2.
The present working hypothesis is that absorption of dietary Cu is related to mRNA expressions of genes involved in Cu uptake and transport of the intestine in fish. To this end, the full-length cDNA sequences of eight Cu uptake related genes, including two isoforms of copper transporter genes (ctr1 and ctr2), three copper chaperone genes (atox1, ccs and cox17), two Cu-ATPase genes (atp7a and atp7b) and divalent metal ion transporter 1 (dmt1), were cloned and characterized in yellow catfish P. fulvidraco, respectively. Their mRNA tissue expression and transcriptional responses to dietborne Cu exposure were investigated. Compared to the corresponding members of mammals, all of these members in P. fulvidraco shared the similar conserved domain structures. Their mRNAs were expressed in a wide range of tissues (including liver, muscle, spleen, brain, gill, intestine, heart and kidney), but at variable levels. In anterior intestine, mRNA levels of ctr1, cox17, dmt1 and atp7a declined with increasing dietary Cu levels. The mRNA levels of ctr2 and mt were the highest for excess dietary Cu group and showed no significant differences between other two treatments. Atox1 mRNA levels were the highest for Cu-deficient group and showed no significant differences between other two treatments. The mRNA levels of ccs were the highest for Cu-deficient group, followed by Cu-excess group and the lowest for adequate-Cu group. In contrast, atp7b mRNA levels were the highest for Cu-excess group and the lowest for adequate Cu group. In the mid-intestine, mRNA levels of ctr1, ctr2, atox1, ccs, cox17, dmt1 and atp7a declined with increasing dietary Cu levels. Atp7b mRNA levels were the lowest for adequate Cu group and showed no significant differences between other two treatments. Mt mRNA levels were the lowest for adequate Cu group and highest for Cu-excess group. For the first time, our study cloned and characterized ctr1, ctr2, atox1, ccs, cox17, atp7a, atp7b and dmt1 genes in P. fulvidraco and determined their tissue-specific expression, and transcriptional responses in the anterior and mid-intestine of yellow catfish under dietborne Cu exposure, which shed new light on the Cu uptake system and help to understand the molecular mechanisms of Cu homeostasis in fish.
目前的工作假说认为，膳食铜的吸收与鱼类肠道中涉及铜摄取和转运的基因的 mRNA 表达有关。为此，本研究分别克隆并鉴定了黄颡鱼（Pelteobagrus fulvidraco）中 8 个铜摄取相关基因的全长 cDNA 序列，包括 2 个铜转运体基因（ctr1 和 ctr2）的 2 个同工型、3 个铜伴侣基因（atox1、ccs 和 cox17）、2 个 Cu-ATPase 基因（atp7a 和 atp7b）和二价金属离子转运体 1（dmt1）。研究了它们在不同组织中的 mRNA 表达谱和对饮食铜暴露的转录响应。与哺乳动物的相应成员相比，这些黄颡鱼成员都具有相似的保守结构域。它们的 mRNA 在广泛的组织（包括肝脏、肌肉、脾脏、大脑、鳃、肠、心脏和肾脏）中表达，但水平不同。在前肠中，随着膳食铜水平的增加，ctr1、cox17、dmt1 和 atp7a 的 mRNA 水平下降。ctr2 和 mt 的 mRNA 水平在过量膳食铜组中最高，与其他两组无显著差异。atox1 的 mRNA 水平在铜缺乏组中最高，与其他两组无显著差异。ccs 的 mRNA 水平在铜缺乏组中最高，其次是铜过量组，在铜适量组中最低。相反，atp7b 的 mRNA 水平在铜过量组中最高，在铜适量组中最低。在中肠中，随着膳食铜水平的增加，ctr1、ctr2、atox1、ccs、cox17、dmt1 和 atp7a 的 mRNA 水平下降。atp7b 的 mRNA 水平在铜适量组中最低，与其他两组无显著差异。mt 的 mRNA 水平在铜适量组中最低，在铜过量组中最高。本研究首次在黄颡鱼中克隆和鉴定了 ctr1、ctr2、atox1、ccs、cox17、atp7a、atp7b 和 dmt1 基因，并确定了它们在鱼体铜暴露条件下前肠和中肠的组织特异性表达和转录响应，为铜摄取系统提供了新的认识，并有助于理解鱼类铜稳态的分子机制。
