Integration of a Galdieria plasma membrane sugar transporter enables heterotrophic growth of the obligate photoautotrophic red alga Cynanidioschyzon merolae
Integration of a Galdieria plasma membrane sugar transporter enables heterotrophic growth of the obligate photoautotrophic red alga Cynanidioschyzon merolae
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DOI：
10.1002/pld3.134
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发表时间：
2019-04-01
期刊：
影响因子：
3
通讯作者：
Miyagishima, Shin-ya
中科院分区：
文献类型：
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作者：
Fujiwara, Takayuki;Hirooka, Shunsuke;Miyagishima, Shin-ya
单细胞嗜热嗜酸红蓝细菌Cyanidioschyzonmerolae是一种新兴的光合真核生物模式生物。其相对简单的基因组（16.5 Mbp）具有非常低的遗传冗余度，其细胞结构具有一个叶绿体，叶绿体，过氧化物酶体和其他细胞器，促进了研究。此外，该基因组在遗传上是易处理的，并且核和叶绿体基因组可以通过经由同源重组的转基因整合来修饰。最近的研究试图阐明这种植物光系统的结构和功能。然而，由于该菌是专性自养生物，很难获得光合缺陷突变体用于分子遗传学研究。为了解决这个问题，在C。在merolae中，我们表达了一个来自Galdieria sulphuraria的质膜糖转运蛋白GsSPT 1，它是C. merolae和能够异养生长。异源表达的GsSPT 1定位于质膜。GsSPT 1使能C。merolae的生长mixotrophically和异养，其中细胞生长在黑暗中与葡萄糖或在光与光合作用抑制剂3-（3，4-二氯苯基）-1，1-二甲基脲（DCMU）和葡萄糖。当GsSPT 1转基因在C.通过URA（Cm-Gs）选择标记，可以使自身及其侧翼转基因植株增殖，GsSPT 1蛋白水平增加，促进了该菌的异养和兼养生长。我们还发现过表达GsSPT 1的C. merolae在光下在含有葡萄糖和DCMU的固化培养基上有效地形成菌落。因此，GsSPT 1过表达将有利于单菌落分离和光合作用缺陷突变体的随机或定点诱变产生的分析。此外，我们的研究结果也支持了质膜糖转运蛋白的存在或缺乏是导致C. merolae和G.硫磺。
The unicellular thermoacidophilic red alga Cyanidioschyzon merolae is an emerging model organism of photosynthetic eukaryotes. Its relatively simple genome (16.5 Mbp) with very low-genetic redundancy and its cellular structure possessing one chloroplast, mitochondrion, peroxisome, and other organelles have facilitated studies. In addition, this alga is genetically tractable, and the nuclear and chloroplast genomes can be modified by integration of transgenes via homologous recombination. Recent studies have attempted to clarify the structure and function of the photosystems of this alga. However, it is difficult to obtain photosynthesis-defective mutants for molecular genetic studies because this organism is an obligate autotroph. To overcome this issue in C. merolae, we expressed a plasma membrane sugar transporter, GsSPT1, from Galdieria sulphuraria, which is an evolutionary relative of C. merolae and capable of heterotrophic growth. The heterologously expressed GsSPT1 localized at the plasma membrane. GsSPT1 enabled C. merolae to grow mixotrophically and heterotrophically, in which cells grew in the dark with glucose or in the light with a photosynthetic inhibitor 3-(3,4-dichlorophenyl)-1,1-dimethylurea (DCMU) and glucose. When the GsSPT1 transgene multiplied on the C. merolae chromosome via the URA(Cm-Gs) selection marker, which can multiply itself and its flanking transgene, GsSPT1 protein level increased and the heterotrophic and mixotrophic growth of the transformant accelerated. We also found that GsSPT1 overexpressing C. merolae efficiently formed colonies on solidified medium under light with glucose and DCMU. Thus, GsSPT1 overexpresser will facilitate single colony isolation and analyses of photosynthesis-deficient mutants produced either by random or site-directed mutagenesis. In addition, our results yielded evidence supporting that the presence or absence of plasma membrane sugar transporters is a major cause of difference in trophic properties between C. merolae and G. sulphuraria.