Max-Planck-Institut für Molekulare Pflanzenphysiologie, Potsdam-Golm, Germany.
Plant Physiol. 2021 Apr 2;185(3):1091-1110. doi: 10.1093/plphys/kiaa106.
De novo fatty acid biosynthesis in plants relies on a prokaryotic-type acetyl-CoA carboxylase (ACCase) that resides in the plastid compartment. The enzyme is composed of four subunits, one of which is encoded in the plastid genome, whereas the other three subunits are encoded by nuclear genes. The plastid gene (accD) encodes the β-carboxyltransferase subunit of ACCase and is essential for cell viability. To facilitate the functional analysis of accD, we pursued a transplastomic knockdown strategy in tobacco (Nicotiana tabacum). By introducing point mutations into the translational start codon of accD, we obtained stable transplastomic lines with altered ACCase activity. Replacement of the standard initiator codon AUG with UUG strongly reduced AccD expression, whereas replacement with GUG had no detectable effects. AccD knockdown mutants displayed reduced ACCase activity, which resulted in changes in the levels of many but not all species of cellular lipids. Limiting fatty acid availability caused a wide range of macroscopic, microscopic, and biochemical phenotypes, including impaired chloroplast division, reduced seed set, and altered storage metabolism. Finally, while the mutants displayed reduced growth under photoautotrophic conditions, they showed exaggerated growth under heterotrophic conditions, thus uncovering an unexpected antagonistic role of AccD activity in autotrophic and heterotrophic growth.
植物从头合成脂肪酸依赖于定位于质体隔室中的原核型乙酰辅酶 A 羧化酶（ACCase）。该酶由四个亚基组成，其中一个亚基由质体基因组编码，而另外三个亚基由核基因编码。质体基因（accD）编码 ACCase 的β-羧基转移酶亚基，对于细胞活力是必需的。为了促进 accD 的功能分析，我们在烟草（Nicotiana tabacum）中采用了质体转化敲低策略。通过在 accD 的翻译起始密码子中引入点突变，我们获得了具有改变的 ACCase 活性的稳定质体转化系。用 UUG 替代标准起始密码子 AUG 强烈降低了 AccD 的表达，而用 GUG 替代则没有可检测到的影响。AccD 敲低突变体表现出 ACCase 活性降低，导致许多但不是所有细胞脂质种类的水平发生变化。脂肪酸可用性的限制导致了广泛的宏观、微观和生化表型，包括叶绿体分裂受损、种子产量降低和储存代谢改变。最后，尽管突变体在光自养条件下的生长受到抑制，但它们在异养条件下表现出过度生长，从而揭示了 AccD 活性在自养和异养生长中的意想不到的拮抗作用。
