Cosson J, White D, Huitorel P, Eddé B, Cibert C, Audebert S, Gagnon C
URA 671 CNRS, Univ. P. et M. Curie, Villefranche-sur-Mer, France.
Cell Motil Cytoskeleton. 1996;35(2):100-12. doi: 10.1002/(SICI)1097-0169(1996)35:2<100::AID-CM3>3.0.CO;2-E.
A panel of monoclonal antibodies (mAbs) has been generated against sea urchin sperm axonemes and selected for their ability to inhibit the motility of sea urchin sperm models. The mAb C9 recognized a 50 kDa protein on blots of sea urchin sperm axonemes. Two-dimensional sodium dodecyl sulfate-polyacrylamide gel electrophoresis revealed that C9 recognized isoforms of beta-tubulin. Low concentrations of C9 (0.1-1.0 microgram/ml) blocked the motility of sea urchin sperm models by decreasing the sliding velocity and frequency of flagellar beating to less than 1 Hz and by modifying the shear angle along the axoneme, especially the distal end. Other antitubulin antibodies had little effect on motility at concentrations 100-fold higher than those effective for C9. The effects on motility were not restricted to flagella of sea urchin spermatozoa. Flagellar beating of the dinoflagellate Oxyrrhis marina was completely blocked by C9 in a manner reminiscent of that of sea urchin sperm flagella. The mAb also inhibited the motility of human spermatozoa and Chlamydomonas reinhardtii. Immunofluorescence techniques revealed that C9 stains the whole axoneme of sea urchin spermatozoa and O. marina flagella together with the cortical network of O. marina cell body. C9 is the first antitubulin antibody reported to interfere with flagellar beat frequency. The observation that this antibody arrests the motility of flagella from sea urchin sperm along with that of dinoflagellate, algae, and human sperm flagella suggests that the epitope recognized by C9 is conserved over a long period of evolution and plays an important role in sperm motility.
已制备出一组针对海胆精子轴丝的单克隆抗体（mAb），并根据其抑制海胆精子模型运动的能力进行筛选。单克隆抗体C9在海胆精子轴丝印迹上识别出一种50 kDa的蛋白质。二维十二烷基硫酸钠-聚丙烯酰胺凝胶电泳显示，C9识别β-微管蛋白的同工型。低浓度的C9（0.1-1.0微克/毫升）通过将鞭毛摆动的滑动速度和频率降低至小于1赫兹，并通过改变沿轴丝尤其是远端的剪切角，来阻断海胆精子模型的运动。其他抗微管蛋白抗体在浓度比C9有效浓度高100倍时对运动几乎没有影响。对运动的影响并不局限于海胆精子的鞭毛。C9完全阻断了双鞭毛虫海洋尖尾藻的鞭毛摆动，其方式类似于海胆精子鞭毛。该单克隆抗体还抑制了人类精子和莱茵衣藻的运动。免疫荧光技术显示，C9可对海胆精子的整个轴丝以及海洋尖尾藻鞭毛进行染色，同时也可对海洋尖尾藻细胞体的皮质网络进行染色。C9是首个被报道可干扰鞭毛摆动频率的抗微管蛋白抗体。该抗体能阻止海胆精子鞭毛以及双鞭毛虫、藻类和人类精子鞭毛的运动，这一观察结果表明，C9识别的表位在长期进化过程中是保守的，并且在精子运动中起着重要作用。
