Sperm cells of Zea mays have a complex complement of mRNAs
Sperm cells of Zea mays have a complex complement of mRNAs
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DOI：
10.1046/j.1365-313x.2003.01761.x
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发表时间：
2003-06-01
期刊：
影响因子：
7.2
通讯作者：
McCormick, S
中科院分区：
文献类型：
--
作者：
Engel, ML;Chaboud, A;McCormick, S
尽管被子植物的双受精作用早在1898年就被发现，但我们对植物中介导配子识别和融合的蛋白质仍然一无所知。由于精子很小，并且嵌在花粉粒的大型营养细胞中，因此来自精子的mRNA在EST数据库中的代表性很差。我们优化了荧光激活细胞分选（FACS），以分离玉米精子无污染的营养细胞质，并构建了cDNA文库。对来自未扩增文库的超过1100个cDNA的测序显示，精子具有多种多样的mRNA互补。大多数转录本都是单基因的，最丰富的转录本只被测序了17次。约8%的序列被预测编码分泌或质膜定位的蛋白质，因此可能介导配子相互作用的候选人。大约8%的序列对应于反转录子。植物精子具有浓缩的染色质，并且被认为是转录不活跃的。我们使用RT-PCR和原位杂交来确定选定的精子mRNA转录的时间。精子转录本编码的蛋白质参与一般细胞功能，在整个花粉发育和更丰富的三细胞花粉比精子细胞，这表明这些转录本也存在于较大的营养细胞。然而，几个转录本，其中编码的蛋白质是最相似的假设拟南芥蛋白质，似乎只存在于成熟花粉内的精细胞，但已经存在于单细胞小孢子。这表明某些转录本可能在花粉发育早期转录，然后分配到精细胞中。
Although double fertilization in angiosperm was discovered in 1898, we still know nothing about the proteins that mediate gamete recognition and fusion in plants. Because sperm are small and embedded within the large vegetative cell of the pollen grain, mRNAs from sperm are poorly represented in EST databases. We optimized fluorescence-activated cell sorting (FACS) in order to isolate Zea mays sperm free of contaminating vegetative cell cytoplasm, and constructed a cDNA library. Sequencing of over 1100 cDNAs from the unamplified library revealed that sperm have a diverse complement of mRNAs. Most transcripts were singletons; the most abundant was sequenced only 17 times. About 8% of the sequences are predicted to encode secreted or plasma membrane-localized proteins and are therefore candidates that might mediate gamete interactions. About 8% of the sequences correspond to retroposons. Plant sperm have condensed chromatin and are thought to be transcriptionally inactive. We used RT-PCR and in situ hybridization to determine when selected sperm mRNAs were transcribed. Sperm transcripts encoding proteins involved in general cell functions were present throughout pollen development and were more abundant in tricellular pollen than in sperm cells, suggesting that these transcripts were also present in the larger vegetative cell. However, several transcripts, which encode proteins that are most similar to hypothetical Arabidopsis proteins, appeared to be present exclusively in the sperm cells inside mature pollen, but were already present in unicellular microspores. This suggests that certain transcripts might be transcribed early during pollen development and later partitioned into the sperm cells.