Microtubule sliding movement in tilapia sperm flagella axoneme is regulated by Ca2+/calmodulin-dependent protein phosphorylation.
Microtubule sliding movement in tilapia sperm flagella axoneme is regulated by Ca2+/calmodulin-dependent protein phosphorylation.
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DOI：
10.1002/cm.20137
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发表时间：
2006-08
影响因子：
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通讯作者：
M. Morita;A. Takemura;A. Nakajima;M. Okuno
中科院分区：
文献类型：
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作者：
M. Morita;A. Takemura;A. Nakajima;M. Okuno
脱膜罗非鱼精子在10~(-6)M Ca~(2+)浓度下复活。当Ca~(2+)浓度从10~(-6)M增加到10~(-5)M时，运动特征发生改变，虽然拍频没有增加，但鞭毛搏动的剪切角和波幅增大，提示代表动力蛋白活性的轴丝微管滑动速度随着Ca~(2+)浓度的增加而增加。因此，由于动力蛋白活性的增强，可能是由于Ca(2+)与鞭毛蛋白结合而激活鞭毛运动。用~(45)Ca叠加法和免疫学方法检测到一种钙结合蛋白(18 kDa，等电点4.0)，即钙调蛋白(CaM)。CaM拮抗剂W-7抑制了复活率和游泳速度，表明18 kDa的钙结合蛋白是CaM，CaM对鞭毛运动有调节作用。免疫组化检测到CaMKIV在轴丝低离子提取物部分和轴丝残留物中均为一条48 kDa的单一条带，表明CaMKIV与轴丝中不同的位置结合。CaMKIV可能以钙/钙调素依赖的方式磷酸化轴丝蛋白，从而调节动力蛋白的活性。A(32)P摄取显示48、75、120、200、250、380和400 kDa的蛋白被磷酸化，这种磷酸化依赖于Ca(2+)/CaM激酶。在10(-5)M Ca(2+)存在下，蛋白质(380 KDa)被磷酸化。Ca(2+)的增加可能诱导了Ca(2+)/CaM激酶依赖的调节，包括激活/调节鞭毛轴线动力蛋白活性的蛋白磷酸化。
Demembranated euryhaline tilapia Oreochromis mossambicus sperm were reactivated in the presence of concentrations in excess of 10(-6) M Ca(2+). Motility features changed when Ca(2+) concentrations were increased from 10(-6) to 10(-5) M. Although the beat frequency did not increase, the shear angle and wave amplitude of flagellar beating increased, suggesting that the sliding velocity of microtubules in the axoneme, which represents dynein activity, rises with an increase in Ca(2+). Thus, it is possible that Ca(2+) binds to flagellar proteins to activate flagellar motility as a result of the enhanced dynein activity. One Ca(2+)-binding protein (18 kDa, pI 4.0), calmodulin (CaM), was detected by (45)Ca overlay assay and immunologically. A CaM antagonist, W-7, suppressed the reactivation ratio and swimming speed, suggesting that the 18 kDa Ca(2+)-binding protein is CaM and that CaM regulates flagellar motility. CaMKIV was detected immunologically as a single 48 kDa band in both the fraction of low ion extract of the axoneme and the remnant of the axoneme, suggesting that CaMKIV binds to distinct positions in the axoneme. It is possible that CaMKIV phosphorylates the axonemal proteins in a Ca(2+)/CaM-dependent manner for regulating the dynein activity. A (32)P-uptake in the axoneme showed that 48, 75, 120, 200, 250, 380, and 400 kDa proteins were phosphorylated in a Ca(2+)/CaM kinase-dependent manner. Proteins (380 kDa) were phosphorylated in the presence of 10(-5) M Ca(2+). It is possible that an increase in Ca(2+) induces Ca(2+)/CaM kinase-dependent regulation, including protein phosphorylation for activation/regulation of dynein activity in flagellar axoneme.