Regulatory inversion in NAC networks steers the timing of age-dependent cell death in plants [Nanostring]. Arabidopsis thaliana

Aging is a time-dependent biological phenomenon governed by complex networks of regulatory components and their transitions over lifetime. Yet, there have been limited efforts to pin down age-associated networks and map their dynamic characteristics onto aging phenotypes. Here, we built time-course genetic regulatory networks of NAM/ATAF/CUC (NAC) transcription factors during the course of leaf aging in Arabidopsis, using causal regulatory relationships among NACs identified from mutants of 49 aging-associated NACs. These temporal networks revealed a regulatory inversion from activating to repressive regulatory modes at a pre-senescent stage. The inversion was governed by three hub NACs, and their mutants conferred earlier aging with altered expression of reactive oxygen species and salicylic acid response genes. Overexpression of the hub NACs delayed the regulatory inversion, rendering delayed age-dependent cell death. We conclude that the regulatory inversion in NAC networks at a pre-senescent stage directs when age-dependent cell death should proceed in plants. Overall design: We measured mRNA expression of 49 NACs in wild type (col-0) and 49 knockout mutant lines of NACs (duplicate for each mutant) at 4 stages (14, 18, 22, and 26 days of leaf age) 434 samples consist of: 34 wild type samples (=10, 8, 8 and 8 replicates for the four time points, respectively) 16 samples of anac092 mutant, one of the ANAC gene (=4 replicates for each of four time points) 384 samples of 48 anac mutants including at3g12910 and ntl9 (=2 replicates for each of four time points).

衰老是一种随时间进程发生的生物学现象，受复杂的调控组分网络及其在生命周期中的动态转变所支配。然而，当前针对衰老相关调控网络的研究仍较为匮乏，且鲜有研究能将其动态特征精准映射至衰老表型。本研究以拟南芥（Arabidopsis）叶片衰老过程中的NAM/ATAF/CUC（NAC）转录因子为研究对象，基于49个衰老相关NAC突变体所鉴定得到的NAC间因果调控关系，构建了其时间序列遗传调控网络。这些时序调控网络揭示，在衰老前期存在从激活调控模式向抑制调控模式的转换。该调控转换受三个核心枢纽NAC转录因子所调控，它们的突变会导致植株早衰，并伴随活性氧（reactive oxygen species）与水杨酸（salicylic acid）响应基因的表达谱发生改变。过表达这三个枢纽NAC转录因子可延缓该调控转换进程，进而使年龄依赖型细胞死亡延迟发生。综上，拟南芥叶片衰老前期NAC调控网络的转换方向，决定了植物年龄依赖型细胞死亡的发生时序。整体实验设计：我们在叶片衰老的4个阶段（叶龄14、18、22和26天），检测了野生型（Col-0）以及49个NAC敲除突变体株系的49个NAC基因的mRNA表达水平（每个突变体设置2次生物学重复）。本次实验共包含434个样本：其中野生型样本34个（对应上述4个时间点的重复数分别为10、8、8和8）；ANAC基因家族成员anac092突变体样本16个（4个时间点各设4次重复）；其余48个anac突变体（包含at3g12910与ntl9）的样本共384个（每个突变体对应4个时间点各2次重复）。