Wong P W, Pessah I N
Department of Molecular Biosciences, School of Veterinary Medicine, University of California, Davis 95616, USA.
Mol Pharmacol. 1996 Apr;49(4):740-51.
We investigated a novel molecular mechanism by which polychlorinated biphenyls (PCBs) alter microsomal Ca2+ transport with sarcoplasmic reticulum (SR) membranes isolated from skeletal and cardiac muscles. Aroclors with an intermediate weight percent of chlorine enhance by >6-fold the binding of 1 nM[3H]ryanodine to its conformationally sensitive site on the SR Ca2+ -release channel [i.e., ryanodine receptor (RyR)] with high potency (EC50=1.4 microM), whereas Aroclors with either high or low chlorine composition show little activity. Structure-activity studies with selected pentachlorobiphenyl congeners reveal a stringent structural requirement for chlorine substitution at the ortho-positions, with 2,2',3,5',6-pentachlorobiphenyl having the highest potency toward skeletal and cardiac isoforms of RyR (EC50=330 nM and 2 microM, respectively). In contrast, 3,3',4,4',5-pentachlorobiphenyl does not enhance ryanodine binding, suggesting that noncoplanarity of the biphenyl rings is required for channel activation. However, 2,2',4,6,6'-pentachlorobiphenyl is significantly less active toward RyR, suggesting that some degree of rotation about the biphenyl bond is required. 2,2',3,5',6-Pentachlorobiphenyl induces a dose-dependent release of Ca2+ from actively loaded SR vesicles with a maximum rate of 1.2 micromol mg-1 min-1 (EC50=1 microM), whereas 3,3',4,4',5-pentachlorobiphenyl (< / = microM) does not alter Ca2+ transport. The mechanism of PCB-induced channel activation involves a significant decrease in the inhibitory potency of Ca2+ and Mg2+ (20-fold and 100-fold, respectively). Neither 2,2',3,5',6- nor 3,3',4,4',5-pentachlorobiphenyl (< / = 10 microM) alters the activity of the skeletal isoform of sarcoplasmic/endoplasmic reticulum Ca2+ -ATPase or the cardiac isoform of sarcoplasmic/endoplasmic reticulum Ca2+ -ATPase, and PCB-induced Ca2+ release can be fully blocked by either microM ryanodine or ruthenium red. These results are the first to demonstrate a selective ryanodine receptor-mediated mechanism by which ortho-substituted PCBs alter microsomal Ca2+ transport and may have toxicological relevance.
我们研究了一种新的分子机制，通过该机制多氯联苯（PCBs）可改变从骨骼肌和心肌分离的肌浆网（SR）膜微粒体的Ca2+转运。氯含量中等的Aroclors能以高效力（EC50 = 1.4 microM）使1 nM [3H]ryanodine与其在SR Ca2+释放通道[即ryanodine受体（RyR）]上的构象敏感位点的结合增强>6倍，而氯组成高或低的Aroclors几乎没有活性。对选定的五氯联苯同系物进行的构效关系研究表明，邻位氯取代有严格的结构要求，2,2',3,5',6 - 五氯联苯对RyR的骨骼肌和心脏同工型具有最高效力（EC50分别为330 nM和2 microM）。相比之下，3,3',4,4',5 - 五氯联苯不会增强ryanodine结合，这表明联苯环的非共面性是通道激活所必需的。然而，2,2',4,6,6' - 五氯联苯对RyR的活性明显较低，这表明需要围绕联苯键有一定程度的旋转。2,2',3,5',6 - 五氯联苯能诱导从主动加载的SR囊泡中剂量依赖性释放Ca2+，最大速率为1.2 micromol mg-1 min-1（EC50 = 1 microM），而3,3',4,4',5 - 五氯联苯（≤ microM）不会改变Ca2+转运。PCB诱导通道激活的机制涉及Ca2+和Mg2+抑制效力的显著降低（分别降低20倍和100倍）。2,2',3,5',6 - 五氯联苯和3,3',4,4',5 - 五氯联苯（≤10 microM）均不会改变肌浆网/内质网Ca2+ - ATP酶的骨骼肌同工型或肌浆网/内质网Ca2+ - ATP酶的心脏同工型的活性，并且PCB诱导的Ca2+释放可被 microM ryanodine或钌红完全阻断。这些结果首次证明了一种选择性的ryanodine受体介导的机制，通过该机制邻位取代的PCBs可改变微粒体Ca2+转运，并且可能具有毒理学相关性。
