Ashley C C, Lea T J, Hoar P E, Kerrick W G, Strang P F, Potter J D
Friday Harbor Laboratories, WA 98250.
J Muscle Res Cell Motil. 1991 Dec;12(6):532-42. doi: 10.1007/BF01738441.
Two isoforms of troponin C (BTnC1 and BTnC2) from the striated muscle of the arthropod Balanus nubilus Darwin (giant barnacle) have been purified (Potter et al., 1987; Collins et al., 1991). Both isoforms were present in all of the white striated muscle fibres studied but not in the red fibres. The ratio of BTnC2 to BTnC1 in different fibre types varied between 3:1 and 1:1. Both forms of TnC could be readily extracted from myofibrillar bundles of barnacle muscle in low ionic strength EDTA solutions, reducing force activation to less than 10%. Both forms either separately or together reassociated with the TnC-depleted fibres in a relaxing (LR) solution (pCa greater than 8.0, [Mg2+] free = 1 mM, I = 0.15 M), and the reconstituted fibres could be subsequently activated in contraction (LA) solution (pCa = less than 3.8, [Mg2+] free = 1 mM, I = 0.15 M). The dissociation of BTnC 1 + 2 is blocked in low ionic strength solutions containing Mg2+ (greater than or equal to 10 mM). The two isoforms of crayfish TnC (CrTnC1 and CrTnC2) were also found to be equivalent to the barnacle TnCs in their ability to reactivate TnC-depleted barnacle myofibrillar bundles. Similar experiments using rabbit skeletal muscle TnC (STnC) (I = 0.15 M) in BTnC-depleted myofibrillar bundles of barnacle showed considerable variability. STnC could associate, although weakly, with the depleted bundles in either LR or LA, and force could be partially restored. In neither situation was it as effective as either BTnC or CrTnC. Interestingly, bovine cardiac TnC (CTnC), although it did not associate at pCa greater than 7.0, did associate and effectively activate force at pCa less than 3.8, but dissociated on return to pCa greater than 7.0 (LR). Neither barnacle TnC isoform associated with TnC-depleted skinned fibres from rabbit skeletal muscle at pCa greater than 7.0, but did associate and activate these fibres at pCa less than 3.8. Once these fibres were returned to LR and then placed in LA at pCa 3.8 all BTnC-restored force was lost, indicating a dissociation of BTnC once the Ca2+ is lowered, as observed with CTnC in barnacle myofibrillar bundles. Finally, the inhibitory effect of BTnI on force and the absence of an effect of calmodulin, trifluoperazine or ATP-gamma-S on force were all taken as evidence for a thin filament regulated Ca2+ control system.
节肢动物巴氏藤壶（Balanus nubilus Darwin，巨型藤壶）横纹肌中的两种肌钙蛋白C同工型（BTnC1和BTnC2）已被纯化（Potter等人，1987年；Collins等人，1991年）。在所有研究的白色横纹肌纤维中均存在这两种同工型，但在红色纤维中不存在。不同纤维类型中BTnC2与BTnC1的比例在3:1至1:1之间变化。两种形式的肌钙蛋白C（TnC）都可以在低离子强度的EDTA溶液中很容易地从藤壶肌肉的肌原纤维束中提取出来，使力激活降低到10%以下。两种形式单独或一起在松弛（LR）溶液（pCa大于8.0，游离[Mg2+]=1 mM，I = 0.15 M）中与耗尽TnC的纤维重新结合，并且重构的纤维随后可以在收缩（LA）溶液（pCa小于3.8，游离[Mg2+]=1 mM，I = 0.15 M）中被激活。BTnC 1 + 2的解离在含有Mg2+（大于或等于10 mM）的低离子强度溶液中被阻断。小龙虾TnC的两种同工型（CrTnC1和CrTnC2）在重新激活耗尽TnC的藤壶肌原纤维束的能力方面也被发现与藤壶TnC相当。在藤壶耗尽BTnC的肌原纤维束中使用兔骨骼肌TnC（STnC）（I = 0.15 M）进行的类似实验显示出相当大的变异性。STnC可以结合，尽管很弱，在LR或LA中与耗尽的纤维束结合，并且力可以部分恢复。在这两种情况下，它都不如BTnC或CrTnC有效。有趣的是，牛心肌TnC（CTnC）虽然在pCa大于7.0时不结合，但在pCa小于3.8时确实结合并有效激活力，但在回到pCa大于7.0（LR）时解离。在pCa大于7.0时，两种藤壶TnC同工型都不与兔骨骼肌去表皮纤维中耗尽TnC的纤维结合，但在pCa小于3.8时确实结合并激活这些纤维。一旦这些纤维回到LR，然后在pCa 3.8时置于LA中，所有BTnC恢复的力都丧失了，这表明一旦Ca2+降低，BTnC就会解离，就像在藤壶肌原纤维束中观察到的CTnC一样。最后，BTnI对力的抑制作用以及钙调蛋白、三氟拉嗪或ATP-γ-S对力没有影响都被视为细丝调节Ca2+控制系统的证据。
