Kubo Takeaki, Abe Jun, Oyamada Takeshi, Ohnishi Makoto, Fukuzawa Hideya, Matsuda Yoshihiro, Saito Tatsuaki
Department of Applied Science, Faculty of Science, Okayama University of Science, Okayama, 700-0005 Japan.
Plant Cell Physiol. 2008 Jun;49(6):981-93. doi: 10.1093/pcp/pcn076. Epub 2008 May 17.
When mating type plus and minus gametes of Chlamydomonas are mixed, they agglutinate with each other via their flagella, fuse, then initiate the zygote formation program which includes synthesis of the zygote cell wall, fusion of nuclei and chloroplasts, and the digestion of chloroplast DNA from the minus parent. The mRNAs from gamete and zygote cells was isolated and hybridized to cDNA-macroarray filters both to identify new genes expressed during the mating reaction and the early zygote formation process and to analyze the gene expression programs that underlie these sexual processes. Twenty-one novel genes were identified in this screen, designated as EZY (early zygote expressed) genes. The EZY genes included genes encoding proteins whose function is unknown, and genes encoding proteins that appear to be involved in processes such as cell wall synthesis, gene expression, intracellular trafficking or secretion, and vesicular transport in zygotic cells. All of the EZY genes were strongly induced within 1 h during the mating process, including early zygote formation. The transcriptional characteristics of EZY genes were analyzed by using the fusion-defective mutant fus mt(+). Among the EZY genes, 12 genes were not activated in fusion-defective conditions, suggesting that cell fusion is required for their expression. The remaining nine that were transcribed in fusion-defective fus matings were also inducible by cell wall removal in either vegetative or gametic cells, indicating that these genes were induced only indirectly by the cAMP signaling pathway initiated by flagellar agglutination as a result of mating-induced cell wall loss.
当衣藻的正交配型和负交配型配子混合时，它们会通过鞭毛相互凝集，融合，然后启动合子形成程序，该程序包括合子细胞壁的合成、细胞核和叶绿体的融合，以及来自负亲本的叶绿体DNA的消化。从配子和合子细胞中分离出mRNA，并与cDNA宏阵列滤膜杂交，以鉴定在交配反应和早期合子形成过程中表达的新基因，并分析这些有性过程背后的基因表达程序。在这个筛选过程中鉴定出了21个新基因，命名为EZY（早期合子表达）基因。EZY基因包括编码功能未知蛋白质的基因，以及编码似乎参与细胞壁合成、基因表达、细胞内运输或分泌以及合子细胞中囊泡运输等过程的蛋白质的基因。所有EZY基因在交配过程（包括早期合子形成）的1小时内都被强烈诱导。通过使用融合缺陷型突变体fus mt(+)分析了EZY基因的转录特征。在EZY基因中，有12个基因在融合缺陷条件下未被激活，这表明它们的表达需要细胞融合。其余9个在融合缺陷型fus交配中被转录的基因，在营养细胞或配子细胞中通过去除细胞壁也可被诱导，这表明这些基因只是由于交配诱导的细胞壁丢失而由鞭毛凝集引发的cAMP信号通路间接诱导的。
