Overexpression of <i>LtKNOX1</i> from <i>Lilium tsingtauense</i> in <i>Nicotiana benthamiana</i> affects the development of leaf morphology

Leaves are the main vegetative organs of the aboveground part of plants and play an important role in plant morphogenesis. <i>KNOTTED-LIKE HOMEOBOX</i> (<i>KNOX</i>) plays a crucial role in regulating leaf cell fate and maintaining leaf development. In this study, we analyzed <i>LtKNOX1</i> from <i>Lilium tsingtauense</i> and illustrated its function in transgenic plants. Tissue-specific expression analysis indicated that <i>LtKNOX1</i> was highly expressed in stems, young flower buds, and shoot apical meristems (SAMs). Ectopic overexpression of <i>LtKNOX1</i> in <i>Nicotiana benthamiana</i> suggested that transformants with mild phenotypes were characterized by foliar wrinkles and mildly curled leaves; transformants with intermediate phenotypes showed severely crimped blades and narrow leaf angles, and the most severe phenotypes lacked normal SAMs and leaves. Moreover, the expression levels of genes involved in the regulation of KNOX in transgenic plants were detected, including <i>ASYMMETRIC LEAVES1, PIN-FORMED 1, GA20-oxidase, CUP-SHAPED COTYLEDON 2, CLAVATA 1</i> and <i>WUSCHEL</i>(<i>WUS</i>), and the expression of other genes were down-regulated except <i>WUS</i>. This study contributes to our understanding of the <i>LtKNOX1</i> function.

叶片是植物地上部分的主要营养器官，在植物形态建成中发挥重要作用。类结状同源盒基因（KNOTTED-LIKE HOMEOBOX，简称KNOX）在调控叶片细胞命运与维持叶片发育方面发挥关键作用。本研究对青岛百合（Lilium tsingtauense）中的LtKNOX1基因进行了分析，并阐明了其在转基因植株中的功能。组织特异性表达分析结果显示，LtKNOX1在茎、幼花蕾以及茎尖分生组织（shoot apical meristems，SAMs）中高表达。将LtKNOX1在本氏烟草（Nicotiana benthamiana）中进行异位过表达后发现：表型轻微的转化植株表现为叶片褶皱、轻度卷曲；表型中等的转化植株叶片严重皱缩、叶夹角变小；而表型最严重的转化植株则缺失正常的茎尖分生组织与叶片。此外，本研究检测了转基因植株中参与KNOX调控通路的相关基因的表达水平，包括不对称叶片1（ASYMMETRIC LEAVES1）、PIN-FORMED 1（PIN-FORMED 1）、赤霉素20-氧化酶（GA20-oxidase）、杯状子叶2（CUP-SHAPED COTYLEDON 2）、CLAVATA1（CLAVATA1）以及WUSCHEL（WUS）；结果显示，除WUS外其余基因的表达量均出现下调。本研究有助于深化对LtKNOX1基因功能的认知。