Margis-Pinheiro M, Martin C, Didierjean L, Burkard G
Institut de Biologie Moléculaire des Plantes du CNRS, Strasbourg, France.
Plant Mol Biol. 1993 Jul;22(4):659-68. doi: 10.1007/BF00047406.
Three chitinases have been shown previously to be induced upon various stresses of bean leaves. Time course studies of mRNA accumulation of two of them (P3- and P4-chitinases) have been studied upon virus infection, mercuric chloride treatment and UV irradiation. In alfalfa mosaic virus (AlMV)-infected plants both mRNAs, absent in uninfected bean leaves, become detectable 36 h after inoculation. A maximum level of mRNAs is reached 84 h after inoculation and, whereas the amount of P3-ch mRNA decreases soon after having reached the maximum, the amount of P4-ch mRNA remains at high levels for several days. In mercuric chloride-treated leaves P4-ch mRNA becomes detectable 1-1.5 h after onset of treatment and a maximum level is observed between 6 h and 24 h after treatment; P3-ch mRNA becomes detectable later than P4-ch mRNA in treated leaves and reaches a maximum as late as 18 h after treatment has been applied. UV light also induces the synthesis of both mRNAs but, here again, important differences are observed in the accumulation rate of the two transcripts. The relative amounts of each mRNA induced by the different stresses have been compared. The most effective inducer of P3-ch mRNA is AlMV. In contrast, mercuric chloride induces P4-ch mRNA more efficiently than AlMV or UV light. We have also determined the complete nucleotide sequence of the cDNA encoding P3-chitinase that has been isolated from a cDNA library by using the cucumber lysozyme-chitinase cDNA as a probe. The 1072 bp P3-ch cDNA encodes a mature protein of 268 amino acid residues and the 25 residue NH2-terminal signal peptide of the precursor. Because of its high structural homology to the cucumber and Arabidopsis acidic chitinases as well as to the N-terminal amino acid sequence of the bifunctional lysozyme-chitinase from P. quinquifolia, bean P3-chitinase can be considered to belong to the class III chitinases. Southern blot analysis of bean genomic DNA revealed that P3-chitinase is encoded by a single gene.
先前已表明，三种几丁质酶在菜豆叶片遭受各种胁迫时会被诱导产生。对其中两种几丁质酶（P3-和P4-几丁质酶）的mRNA积累进行了时间进程研究，研究对象包括病毒感染、氯化汞处理和紫外线照射。在苜蓿花叶病毒（AlMV）感染的植株中，未感染的菜豆叶片中不存在的这两种mRNA，在接种后36小时变得可检测到。接种后84小时达到mRNA的最高水平，然而，P3-ch mRNA在达到最高水平后不久就减少，而P4-ch mRNA的量在几天内保持在高水平。在氯化汞处理的叶片中，P4-ch mRNA在处理开始后1 - 1.5小时变得可检测到，在处理后6小时至24小时之间观察到最高水平；在处理的叶片中，P3-ch mRNA比P4-ch mRNA更晚变得可检测到，并且在施加处理后18小时才达到最高水平。紫外线也诱导这两种mRNA的合成，但在这里，再次观察到两种转录本积累速率的重要差异。比较了不同胁迫诱导的每种mRNA的相对量。P3-ch mRNA最有效的诱导剂是AlMV。相反，氯化汞比AlMV或紫外线更有效地诱导P4-ch mRNA。我们还确定了编码P3-几丁质酶的cDNA的完整核苷酸序列，该序列是通过使用黄瓜溶菌酶-几丁质酶cDNA作为探针从cDNA文库中分离出来的。1072 bp的P3-ch cDNA编码一个由268个氨基酸残基组成的成熟蛋白以及前体的25个残基的NH2-末端信号肽。由于其与黄瓜和拟南芥酸性几丁质酶以及来自五叶参的双功能溶菌酶-几丁质酶的N-末端氨基酸序列具有高度的结构同源性，菜豆P3-几丁质酶可被认为属于III类几丁质酶。对菜豆基因组DNA的Southern印迹分析表明，P3-几丁质酶由单个基因编码。
