Fry S C, Willis S C, Paterson A E
The Edinburgh Cell Wall Group, Institute of Cell and Molecular Biology, The University of Edinburgh, UK.
Planta. 2000 Oct;211(5):679-92. doi: 10.1007/s004250000330.
Maize (Zea mays L.) cell cultures incorporated radioactivity from [14C]cinnamate into hydroxycinnamoyl-CoA derivatives and then into polysaccharide-bound feruloyl residues. Within 5-20 min, the CoA pool had lost its 14C by turnover and little or no further incorporation into polysaccharides then occurred. The system was thus effectively a pulse-chase experiment. Kinetics of radiolabelling of diferulates (also known as dehydrodiferulates) varied with culture age. In young (1-3 d) cultures, polysaccharide-bound [14C]feruloyl- and [14C]diferuloyl residues were both detectable within 1 min of [14C]cinnamate feeding. Thus, feruloyl residues were dimerised < 1 min after their attachment to polysaccharides. For at least the first 2.3 h after [14C]cinnamate feeding, polysaccharide-bound [14C]diferuloyl residues remained almost constant at approximately 7% of the total polysaccharide-bound [14C]ferulate derivatives. Since feruloyl residues are attached to polysaccharides < 1 min after the biosynthesis of the latter, and > 10 min before secretion, the data show that extensive feruloyl coupling occurred intra-protoplasmically. Exogenous H2O2 (1 mM) caused little additional feruloyl coupling; therefore, wall-localised coupling may have been peroxidase-limited. In older (e.g. 4 d) cultures, less intraprotoplasmic coupling occurred: during the first 2.5 h, polysaccharide-bound [14C]diferuloyl residues were a steady 1.4% of the total polysaccharide-bound [14C]ferulate derivatives. In contrast to the situation in younger cultures, exogenous H2O2 induced a rapid 4- to 6-fold increase in all coupling products, indicating that coupling in the walls was H2O2-limited. In both 2- and 4-d-old cultures, polysaccharide-bound 14C-trimers and larger coupling products exceeded [14C]diferulates 3- to 4-fold, but followed similar kinetics. Thus, although all known dimers of ferulate can now be individually quantified, it appears to be trimers and larger products that make the major contribution to cross-linking of wall polysaccharides in cultured maize cells. We argue that feruloyl arabinoxylans that are cross-linked before and after secretion are likely to loosen and tighten the cell wall, respectively. The consequences for the control of cell expansion and for the response of cell walls to an oxidative burst are discussed.
玉米（Zea mays L.）细胞培养物将[14C]肉桂酸的放射性掺入到羟基肉桂酰辅酶A衍生物中，然后再掺入到与多糖结合的阿魏酰残基中。在5 - 20分钟内，辅酶A池通过周转失去了其14C，随后很少或几乎没有进一步掺入到多糖中。因此，该系统实际上是一个脉冲追踪实验。二阿魏酸酯（也称为脱氢二阿魏酸酯）的放射性标记动力学随培养时间而变化。在年轻（1 - 3天）的培养物中，在加入[14C]肉桂酸1分钟内，就可检测到与多糖结合的[14C]阿魏酰和[14C]二阿魏酰残基。因此，阿魏酰残基在附着到多糖后不到1分钟就发生了二聚化。在加入[14C]肉桂酸后的至少前2.3小时内，与多糖结合的[14C]二阿魏酰残基几乎保持恒定，约占与多糖结合的[14C]阿魏酸衍生物总量的7%。由于阿魏酰残基在多糖生物合成后不到1分钟就附着到多糖上，且在分泌前10多分钟就已附着，数据表明大量的阿魏酰偶联发生在细胞质内。外源H2O2（1 mM）几乎没有引起额外的阿魏酰偶联；因此，细胞壁定位的偶联可能受过氧化物酶限制。在较老（如4天）的培养物中，细胞质内的偶联较少：在最初的2.5小时内，与多糖结合的[14C]二阿魏酰残基稳定在与多糖结合的[14C]阿魏酸衍生物总量的1.4%。与年轻培养物的情况相反，外源H2O2诱导所有偶联产物迅速增加4至6倍，表明细胞壁中的偶联受H2O2限制。在2天和4天龄的培养物中，与多糖结合的14C - 三聚体和更大的偶联产物比[14C]二阿魏酸酯高出3至4倍，但遵循相似的动力学。因此，尽管现在所有已知的阿魏酸二聚体都可以单独定量，但似乎是三聚体和更大的产物对培养的玉米细胞中细胞壁多糖的交联起主要作用。我们认为，在分泌前后交联的阿魏酰阿拉伯木聚糖可能分别使细胞壁松弛和收紧。文中讨论了其对细胞扩展控制以及细胞壁对氧化爆发反应的影响。
