GENE-EXPRESSION IN TOBACCO LOW-NICOTINE MUTANTS
GENE-EXPRESSION IN TOBACCO LOW-NICOTINE MUTANTS
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DOI：
10.1105/tpc.6.5.723
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发表时间：
1994-05-01
期刊：
影响因子：
11.6
通讯作者：
YAMADA, Y
中科院分区：
文献类型：
--
作者：
HIBI, N;HIGASHIGUCHI, S;YAMADA, Y
两个核基因Nic 1和Nic 2调节烟草中的尼古丁水平。nic 1和nice是Burley 21中的半显性突变，其降低叶片烟碱水平和烟碱途径中多种酶的活性，同时增加培养根中的多胺水平。培养的两个突变的纯合的根被用来分离两个cDNA通过减法杂交，这两个cDNA的转录水平在突变体的根比在野生型的根低得多。A411基因编码与哺乳动物亚精胺合酶具有相当大同源性的41-kD蛋白，而A622基因编码与β-羟脯氨酸还原酶具有高度同源性的35-kD蛋白。当这些基因在大肠杆菌中表达时，A411没有亚精胺合酶活性，但确实显示腐胺N-甲基转移酶活性，这是第一个致力于尼古丁生物合成途径的酶，A622没有显示过氧化氢还原酶活性。甲基转移酶基因和A622基因均在根中优势表达，其在培养根中的表达量受nic突变体的协同作用，其顺序为野生型> nic 2> nic 1> nic 1 nic 2。此外，外源生长素的供应下调这两个基因在培养的烟草根。这些结果表明，Nic 1和Nice是尼古丁生物合成的调控基因。
Two nuclear genes, Nic1 and Nic2, regulate nicotine levels in tobacco. nic1 and nice are semidominant mutations in Burley 21 that reduce leaf nicotine levels and the activities of multiple enzymes in the nicotine pathway and simultaneously increase polyamine levels in cultured roots. Cultured roots homozygous for both mutations were used to isolate two cDNAs by subtraction hybridization; the transcript levels of these two cDNAs were much lower in the mutant roots than in the wild-type roots. The A411 gene encodes a 41-kD protein with considerable homology to mammalian spermidine synthase, whereas the A622 gene encodes a 35-kD protein with high homology to isoflavone reductase. When these genes were expressed in Escherichia coli, A411 had no spermidine synthase activity but did show putrescine N-methyltransferase activity, which is the first enzyme committed to the nicotine biosynthetic pathway, and A622 did not show isoflavone reductase activity. Both the methyltransferase and A622 genes are predominantly expressed in the root, and their expression levels in cultured roots are coordinately decreased by the nic mutations in the order of wild type > nic2 > nic1 > nic1 nic2 Removal of tobacco flower heads and young leaves rapidly and coordinately induced both genes in the root. Further, exogenous supply of auxin down-regulated both genes in cultured tobacco roots. These results suggest that Nic1 and Nice are regulatory genes for nicotine biosynthesis.