Nitric Oxide Sensing in Plants Is Mediated by Proteolytic Control of Group VII ERF Transcription Factors

Abstract

SummaryNitric oxide (NO) is an important signaling compound in prokaryotes and eukaryotes. In plants, NO regulates critical developmental transitions and stress responses. Here, we identify a mechanism for NO sensing that coordinates responses throughout development based on targeted degradation of plant-specific transcriptional regulators, the group VII ethylene response factors (ERFs). We show that the N-end rule pathway of targeted proteolysis targets these proteins for destruction in the presence of NO, and we establish them as critical regulators of diverse NO-regulated processes, including seed germination, stomatal closure, and hypocotyl elongation. Furthermore, we define the molecular mechanism for NO control of germination and crosstalk with abscisic acid (ABA) signaling through ERF-regulated expression of ABSCISIC ACID INSENSITIVE5 (ABI5). Our work demonstrates how NO sensing is integrated across multiple physiological processes by direct modulation of transcription factor stability and identifies group VII ERFs as central hubs for the perception of gaseous signals in plants.

Gene Regulation and Survival under Hypoxia Requires Starch Availability and Metabolism

“…The quintuple erfVII mutant (Marín-de la ) is a null mutant for RAP2.2, RAP2.3, RAP2.12, HRE1, HRE2, and a knockdown for RAP2.2 (Giuntoli et al, 2017b); a line constitutively expressing a version of the RAP2.12 protein that is also stable in air (35S:D-RAP2.12; Licausi et al, 2011); a line expressing GUS under the control of the PCO1 promoter (pPCO1:GUS; Weits et al, 2014); and a line overexpressing HRA1 . The bzip63 mutant and the bZIP63 overexpressor (35S:bZIP63) (Mair et al, 2015), two independent KIN10 overexpressor lines (OE1 KIN10 and OE2 KIN10) (Baena-González et al, 2007); the Arabidopsis SnRK1.1 dominant-negative mutant (SnRK1.1 K48M ; Cho et al, 2016); a line expressing RAP2.3 with an HA tag (35S:RAP2.3-HA; Gibbs et al, 2014).…”

Large-Scale Identification of Gibberellin-Related Transcription Factors Defines Group VII ETHYLENE RESPONSE FACTORS as Functional DELLA Partners

“…For ChIP assays, we used a transgenic line overexpressing an HA-tagged mutant version of the RAP2.3 protein (35S::MA-RAP2.3-HA). This mutant version of RAP2.3 is constitutively stable; it is resistant to N-end rulemediated degradation, because the Cys at position 2 is replaced by the stabilizing residue Ala (Gibbs et al, 2011(Gibbs et al, , 2014. We did not detect in vivo binding of the RAP2.3 to the region containing the GCC-like box in the promoter of At2g41260 and At4g19690 in etiolated seedlings (Supplemental Fig.…”

“…Group VII ERFs are degraded through the N-end rule pathway in response to the simultaneous presence of nitric oxide (NO) and oxygen (Gibbs et al, 2011(Gibbs et al, , 2014Licausi et al, 2011). The role of group VII ERFs as NO and oxygen sensors and the reported NO-dependent accumulation of DELLA proteins in response to light (Lozano-Juste and León, 2010), therefore, suggest that the interaction with DELLAs might be relevant in other physiological contexts as well.…”

“…The fusion of the HA tag at the N terminus of RAP2.3 will prevent degradation through the N-end rule pathway (Gibbs et al, 2014). As shown in Figure 2C, HA-RAP2.3 was efficiently pulled down from extracts of protoplasts by anti-myc antibodies only in the presence of myc-GAI, indicating that both proteins are able to interact in plants as well.…”

“…In other cases, the interaction with the TF occurs in the vicinity of the promoters of certain genes to promote their expression (Lim et al, 2013;Park et al, 2013). RAP2.3 binds in vitro and in vivo to the GCC box (Büttner and Singh, 1997;Yang et al, 2009;Gibbs et al, 2014), and it is able to activate transcription when transiently overexpressed in Arabidopsis leaves (Yang et al, 2009). To test the effect that the interaction with GAI may have on the transcriptional activation ability of RAP2.3, we performed transactivation assays in leaves of Nicotiana benthamiana.…”