Membrane-Bound Transcriptional Activator NTL1 from Rapeseed Positively Modulates Leaf Senescence through Targeting Genes Involved in Reactive Oxygen Species Production and Programmed Cell Death

Leaf senescence is the last stage of leaf development and is determined by various environmental and endogenous signals. Leaf senescence can determine plant productivity and fitness. Transcription factors (TFs) with the transmembrane domain constitute a special group of regulatory proteins that can translocate from the membrane system into nuclei to exert the transcriptional function upon endogenous or exogenous stimuli. Reactive oxygen species (ROSs) play an important role in numerous processes throughout the life cycle of plants including leaf senescence. Leaf senescence is characterized by massive programmed cell death (PCD) and is a type of developmental PCD. The transcriptional regulatory relationships between membrane-bound TFs and leaf senescence remain largely uncharacterized, especially in rapeseed (Brassica napus L.), an important oil crop. Here, we show that BnaNTL1 is a membrane-bound NAC (NAM, ATAF, and CUC) TF, which is predominantly expressed in senescent leaves. Expression of BnaNTL1ΔTM, a form of BnaNTL1 devoid of the transmembrane domain, can induce serious HR-like cell death symptoms and ROS accumulation in cells. Plants overexpressing BnaNTL1ΔTM show earlier leaf senescence compared with the control, accompanied by chlorophyll degradation and electrolyte leakage. Genes involved in ROS production (RbohD), PCD (VPEs and CEP1), and leaf senescence (BFN1) are significantly induced and activated by BnaNTL1ΔTM according to the quantitative reverse transcription PCR (qRT-PCR) analysis and dual luciferase reporter (Dual-LUC) assay. Moreover, electrophoretic mobility shift assay revealed that BnaNTL1 directly bound to the NTLBS elements in promoters of RbohD, γVPE, and BFN1. In conclusion, these results demonstrate that BnaNTL1 positively modulates ROS production and HR-like cell death to induce leaf senescence.