Sreenivas A, Sastry P S
Department of Biochemistry, Indian Institute of Science, Bangalore.
Arch Biochem Biophys. 1994 Jun;311(2):229-34. doi: 10.1006/abbi.1994.1231.
The developing seeds of Actinodaphne hookeri were investigated to delineate their ability to synthesize large amounts of trilaurin. Until 88 days after flowering the embryos contained 71% neutral lipids (NL) and 29% phospholipids (PL) and both these components contained C16:0, C18:0, C18:2, and C18:3 as the major fatty acids (FA). At 102 days after flowering the seeds began to accumulate triacylglycerols (TAG) and to synthesize lauric acid (C12:0). By 165 days after flowering, when the seeds were mature, they contained about 99% NL and 1% PL. At this stage the TAG contained exclusively C12:0, while the PL consisted of long-chain fatty acids (LCFA) only. Leaf lipids in contrast did not contain any C12:0. Experiments on [1-14C]acetate incorporation into developing seed slices showed that at 88 days after flowering only 4% of the label was in TAG, 1% in diacylglycerols (DAG), and 87% in PL. One hundred two days after flowering seeds incorporated only 2% of the label into TAG, 30% into DAG, and 64% into PL. In contrast at 114 days after flowering 71% of the label was incorporated into TAG, 25% into DAG, and only 2% into PL. Analysis of labeled FA revealed that up to 102 days after flowering it was incorporated only into LCFA, whereas at 114 days after flowering it was incorporated exclusively into C12:0. Furthermore, 67% of the label in PL at 114 days after flowering was found to be dilaurylglycerophosphate. Analysis of the label in DAG at this stage showed that it was essentially in dilaurin species. These observations indicate the induction of enzymes of Kennedy pathway for the specific synthesis of trilaurin at about 114 days after flowering. Homogenates of seeds (114 days after flowering) incubated with labeled FA in the presence of glycerol-3-phosphate and coenzymes A and ATP incorporated 84% of C12:0 and 61% of C14:0, but not C16:0, C18:2, and C18:3, into TAG. In contrast the LCFA were incorporated preferentially into PL. It is concluded that, between 102 and 114 days after flowering, a switch occurs in A. hookeri for the synthesis of C12:0 and trilaurin which is tissue specific. Since the seed synthesizes exclusively C12:0 at 114 days after flowering onwards and incorporates specifically into TAG, this system appears to be ideal for identifying the enzymes responsible for medium-chain fatty acid as well as trilaurin synthesis and for exploiting them for genetic engineering.
对滇桂琼楠（Actinodaphne hookeri）发育中的种子进行了研究，以确定其合成大量月桂精的能力。在开花后88天之前，胚中含有71%的中性脂质（NL）和29%的磷脂（PL），这两种成分均以十六烷酸（C16:0）、十八烷酸（C18:0）、十八碳二烯酸（C18:2）和十八碳三烯酸（C18:3）作为主要脂肪酸（FA）。在开花后102天，种子开始积累三酰甘油（TAG）并合成月桂酸（C12:0）。到开花后165天种子成熟时，它们含有约99%的NL和1%的PL。在此阶段，TAG仅含C12:0，而PL仅由长链脂肪酸（LCFA）组成。相比之下，叶片脂质不含任何C12:0。对[1-14C]乙酸掺入发育中种子切片的实验表明，在开花后88天，仅有4%的标记物存在于TAG中，1%存在于二酰甘油（DAG）中，87%存在于PL中。开花后102天，种子仅将2%的标记物掺入TAG，30%掺入DAG，64%掺入PL。相反，在开花后114天，71%的标记物掺入TAG，25%掺入DAG，仅2%掺入PL。对标记FA的分析表明，在开花后102天之前，它仅掺入LCFA，而在开花后114天，它仅掺入C12:0。此外，在开花后114天，PL中67%的标记物被发现是二月桂酰甘油磷酸酯。对该阶段DAG中标记物的分析表明，它主要存在于二月桂精种类中。这些观察结果表明，在开花后约114天诱导了肯尼迪途径的酶以特异性合成月桂精。开花后114天的种子匀浆在甘油-3-磷酸、辅酶A和ATP存在下与标记FA一起孵育，将84%的C12:0和61%的C14:0掺入TAG，但未将C16:0、C18:2和C18:3掺入TAG。相比之下，LCFA优先掺入PL。得出的结论是，在开花后102至114天之间，滇桂琼楠发生了合成C12:0和月桂精的转变，这是组织特异性的。由于种子在开花后114天起仅合成C12:0并特异性掺入TAG，该系统似乎是鉴定负责中链脂肪酸以及月桂精合成的酶并将其用于基因工程的理想选择。
