21-kDa polypeptide, a low-molecular-weight cyclophilin, is released from pollen of higher plants into the extracellular medium in vitro
21-kDa polypeptide, a low-molecular-weight cyclophilin, is released from pollen of higher plants into the extracellular medium in vitro
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DOI：
10.1007/s00425-003-1177-2
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发表时间：
2004-01
期刊：
影响因子：
4.3
通讯作者：
E. Yokota;T. Ohmori;S. Muto;T. Shimmen
中科院分区：
文献类型：
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作者：
E. Yokota;T. Ohmori;S. Muto;T. Shimmen
钙离子对花粉管的伸长和取向起关键作用。我们发现，当百合（Lilium longiflorumThunb）花粉粒在细胞外培养基中特异性释放出大量的21 kda多肽。，在EGTA或低浓度Ca2+存在下孵育。这种现象还取决于pH值和培养基中mgcl2的浓度；增加mgcl2浓度和降低ph可抑制花粉中21-kDa多肽的释放，在释放21-kDa多肽的培养基中抑制花粉粒的萌发。这种抑制是不可逆的；将花粉粒转移到基础培养基中不萌发。免疫电镜显示，该多肽存在于细胞质、营养细胞核和生殖细胞中。当花粉在含有EGTA的培养基中处理时，细胞质中21-kDa多肽的密度显著降低，而营养细胞核和生殖细胞中的21-kDa多肽密度没有显著降低，表明只有细胞质中的21-kDa多肽被释放到细胞外培养基中。21-kDa多肽也存在于其他高等植物的花粉中，如asTradescantia virginianaL。,烟草tabacumL。（被子植物）和日本柳杉。堂。（裸子植物），并在EGTA存在的情况下释放到培养基中。在c的情况下。而在碱性pH高于8.5的条件下，粳稻的花粉会释放出花粉。21-kDa多肽的表达不具有花粉特异性，因为在百合和烟草的营养器官和细胞中也存在与抗21-kDa多肽血清有免疫反应的21-kDa成分。然而，即使在EGTA存在的情况下，21 kda多肽也不会从培养的烟草BY-2细胞释放到细胞外培养基中。从21 kda多肽中提取的两个肽片段的氨基酸序列与低分子量亲环蛋白（CyP）的氨基酸序列吻合良好。抗21 kda多肽血清可识别犊牛胸腺和A431癌细胞中的CyP A。从百合花粉中纯化得到的21 kda多肽片段具有肽酰-脯氨酸-转异构酶活性，该活性被CyPs酶活性抑制剂环孢素A （cyclosporin A, CsA）抑制。从这些结果，我们得出结论，21 kda多肽是一个低分子量的CyP。本研究表明，高等植物花粉中的CyP在不利条件下被释放到细胞外基质中。
Calcium ions play a key role in the elongation and orientation of pollen tubes. We found that significant amounts of 21-kDa polypeptide were specifically released into the extracellular medium when pollen grains of lily,Lilium longiflorumThunb., were incubated in the presence of EGTA or at low concentrations of Ca2+. This phenomenon was also dependent on pH and on the concentrations of MgCl2in the medium; the release of 21-kDa polypeptide from pollen was suppressed by increasing the MgCl2concentration and by lowering pH. Germination of pollen grains was inhibited in the medium into which the 21-kDa polypeptide had been released. This inhibition was irreversible; germination did not occur on transfer of the pollen grains into basal culture medium. Immuno-electron microscopy using an antibody against 21-kDa polypeptide showed that this polypeptide was present in the cytoplasm, vegetative nucleus and generative cell. When the pollen was treated with a medium containing EGTA, the density of 21-kDa polypeptide in the cytoplasm significantly decreased, but its density in vegetative nuclei and the generative cell did not, suggesting that only cytoplasmic 21-kDa polypeptide was released into the extracellular medium. The 21-kDa polypeptide was also present in the pollen of other higher-plant species, such asTradescantia virginianaL.,Nicotiana tabacumL. (angiosperms), andCryptomeria japonicaD. Don. (gymnosperm), and was also released into the medium in the presence of EGTA. In the case ofC. japonica, however, it was released from pollen at alkaline pH above 8.5. The expression of 21-kDa polypeptide was not pollen-specific, because 21-kDa components immunoreactive with the anti-21-kDa polypeptide serum also existed in vegetative organs and cells of lily or tobacco. However, the 21-kDa polypeptide was not released into the extracellular medium from cultured tobacco BY-2 cells, even in the presence of EGTA. Amino acid sequences of two peptide fragments derived from 21-kDa polypeptide matched well those of low-molecular-weight cyclophilin (CyP). The antiserum against 21-kDa polypeptide recognized the CyP A from calf thymus and that in A431 carcinoma cells. The 21-kDa polypeptide fraction purified from lily pollen possessed peptidyl-prolylcis-transisomerase activity, which was suppressed by cyclosporin A (CsA), an inhibitor of enzyme activities of CyPs. From these results, we concluded that the 21-kDa polypeptide is a low-molecular-weight CyP. The present study showed that CyP in the pollen of higher plants is released into the extracellular matrix under unfavorable conditions.