Genome-wide identification and analysis of the PAL genes from the orchids Apostasia shenzhenica, Dendrobium catenatum and Phalaenopsis equestris

Phenylalanine ammonia-lyase (PAL) is a key gateway enzyme that connects the phenylpropanoid pathway to primary metabolism. The phenylpropanoid pathway plays a vital role in the growth and environmental adaptation of many plants leading to the production of valuable bioactive compounds with industrial and medical applications. In the present study, nine putative PAL genes from three orchids were identified; five in Apostasia shenzhenica and two each in Dendrobium catenatum and Phalaenopsis equestris. Eighteen motifs and four major conserved functional domains were identified as reported in PAL proteins of other species. All the nine PALs were stable based on their computed physicochemical properties and localized in the cytoplasm. The three-dimensional structures of PALs revealed a homo-tetrameric structure consisting of four identical subunits. A total of 21 cis-regulatory elements with known functions were identified from the promoter regions of all PALs which are responsible for various plant responses to light, stress and growth regulators like auxins, gibberellins and abscisic acid. Phylogenetic analysis showed that the studied PAL proteins clustered in two major clades (clade I and II), placing dicot and monocot PALs in two separate monophyletic clades. In silico gene expression of the identified PALs in different vegetative and reproductive tissues revealed the differential expressions based on tissue type and disclosed that the expression of PAL genes was upregulated in all the tissues examined with an exception of PePAL leaf samples where no expression was detected, however, the same being highly expressed in reproductive tissues (PePAL1-labellum; PePAL2-sepal). In case of AsPALs, the expression was found to be highest in reproductive tissues (AsPAL4-maximum in inflorescence). On the other hand, the expression of DcPALs was found to be highest in vegetative tissues (DcPAL2-maximum in root). Based on the medicinal importance of orchids and the significant role of PAL genes in synthesis of bioactive compounds, the functional characterization of PAL genes can be further exploited in genetic improvement of medicinal orchids. Communicated by Ramaswamy H. Sarma

苯丙氨酸解氨酶（Phenylalanine ammonia-lyase, PAL）是连接苯丙烷途径与初生代谢的关键门户酶。苯丙烷途径在诸多植物的生长发育与环境适应过程中发挥至关重要的作用，可合成兼具工业与医学应用价值的珍贵生物活性化合物。本研究从3种兰科植物中鉴定出9个推定PAL基因：深圳拟兰（Apostasia shenzhenica）中包含5个，铁皮石斛（Dendrobium catenatum）与美花蝴蝶兰（Phalaenopsis equestris）各含2个。与其他物种PAL蛋白的研究报道一致，本研究共鉴定到18个基序与4个主要保守功能结构域。基于计算得到的理化性质，所有9个PAL蛋白均为稳定蛋白，且定位于细胞质中。PAL蛋白的三维结构显示其为同源四聚体，由4个完全相同的亚基构成。从所有PAL基因的启动子区域共鉴定出21个功能明确的顺式调控元件（cis-regulatory elements），这些元件可调控植物对光照、胁迫以及生长素、赤霉素、脱落酸等生长调节剂的多种响应。系统发育分析显示，本研究涉及的PAL蛋白可分为两大主要进化支（进化支I与进化支II），双子叶植物与单子叶植物的PAL蛋白分别聚类于两个独立的单系进化支中。对鉴定得到的PAL基因在不同营养与生殖组织中的计算机模拟基因表达分析显示，其表达量因组织类型而异；所有检测组织中的PAL基因表达均被上调，仅美花蝴蝶兰的PePAL叶片样本未检测到表达，且该类基因在生殖组织中高表达：PePAL1在唇瓣中表达量最高，PePAL2在花萼中表达量最高。对于深圳拟兰的PAL基因（AsPALs），其在生殖组织中的表达量最高，其中AsPAL4在花序中表达量达到峰值。而铁皮石斛的PAL基因（DcPALs）则在营养组织中表达量最高，其中DcPAL2在根中表达量最高。鉴于兰科植物的药用价值以及PAL基因在生物活性化合物合成中的关键作用，PAL基因的功能表征可进一步应用于药用兰科植物的遗传改良。本文由Ramaswamy H. Sarma提交