Identification of the Arabidopsis palmitoyl-monogalactosyldiacylglycerol Δ7-desaturase gene FAD5, and effects of plastidial retargeting of Arabidopsis desaturases on the fad5 mutant phenotype
Identification of the Arabidopsis palmitoyl-monogalactosyldiacylglycerol Δ7-desaturase gene FAD5, and effects of plastidial retargeting of Arabidopsis desaturases on the fad5 mutant phenotype
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DOI：
10.1104/pp.104.052951
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发表时间：
2004-12-01
期刊：
影响因子：
7.4
通讯作者：
Shanklin, J
中科院分区：
文献类型：
--
作者：
Heilmann, I;Mekhedov, S;Shanklin, J
十六烷酸7，10，13-三烯酸(16：3 Delta(7，10，13))是拟南芥中含量最丰富的脂肪酸之一，也是类囊体膜的功能成分，是单半乳糖二酰甘油的sn-2酯。拟南芥FAD5突变体缺乏胞质棕榈酰单半乳糖二酰甘油Delta7-去饱和酶FAD5的活性，其生化特征是缺少16：3的Delta(7，10，13)，生理特征是叶绿素含量减少，光抑制后恢复速率降低。虽然FAD5突变已被定位，但FAD5基因并未被明确鉴定，通过FAD5突变表型的互补来进行正式的功能鉴定尚未见报道。从FAD5植物中克隆了两个候选基因(At3g15850和At3g15870)，它们被预测编码Fad5染色体上的叶绿体靶向去饱和酶。在At3g15850(ADS3)中发现了一个将密码子TGG(Trp-98)变为TGA(STOP)的无义突变，而FAD5的At3g15870等位基因与野生型相同(更正了已发表的野生型基因组At3g15870序列中的一个测序错误)。野生型AT3g15850的基因组克隆或cDNA的表达赋予FAD5植物合成163 Delta(7，10，13)和恢复叶片叶绿素含量的能力。携带T-DNA插入AT3g15870的拟南芥具有16：3 Delta(7，10，13)和叶绿素的野生型水平。总而言之，这些数据正式证明了AT3g15850是FAD5。有趣的是，当拟南芥脱饱和酶(ADS)家族的胞外型Delta9-去饱和酶表达为与胞质转运肽的融合时，FAD5表型部分互补。叶片16：3 Delta(7，10，13)水平与叶绿素含量之间的密切关系表明，胞质脂肪酸去饱和酶在类囊体体的形成中发挥了作用。
Hexadeca 7,10,13-trienoic acid (16:3Delta(7,10,13)) is one of the most abundant fatty acids in Arabidopsis (Arabidopsis thaliana) and a functional component of thylakoid membranes, where it is found as an sn-2 ester of monogalactosyldiacylglycerol. The Arabidopsis fad5 mutant lacks activity of the plastidial palmitoyl-monogalactosyldiacylglyceroI Delta7-desaturase FAD5, and is characterized biochemically by the absence of 16:3Delta(7,10,13) and physiologically by reduced chlorophyll content and a reduced recovery rate after photoinhibition. While the fad5 mutation has been mapped, the FAD5 gene was not unambiguously identified, and a formal functional characterization by complementation of fad5 mutant phenotypes has not been reported. Two candidate genes (At3g15850 and At3g15870) predicted to encode plastid-targeted desaturases at the fad5 chromosomal locus were cloned from fad5 plants and sequenced. A nonsense mutation changing codon TGG (Trp-98) into TGA (stop) was identified in At3g15850 (ADS3), whereas the fad5 At3g15870 allele was identical to wild type (after correction of a sequencing error in the published wild-type genomic At3g15870 sequence). Expression of a genomic clone or cDNA for wild-type At3g15850 conferred on fad5 plants the ability to synthesize 163Delta(7,10,13) and restored leaf chlorophyll content. Arabidopsis carrying a T-DNA insertion in At3g15870 had wild-type levels of both 16:3Delta(7,10,13) and chlorophyll. Together, these data formally prove that At3g15850 is FAD5. Interestingly, the fad5 phenotype was partially complemented when extraplastidial Delta9-desaturases of the Arabidopsis desaturase (ADS) family were expressed as fusions with a plastidial transit peptide. Tight correlation between leaf 16:3Delta(7,10,13) levels and chlorophyll content suggests a role for plastidial fatty acid desaturases in thylakoid formation.