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.

Screening of a genomic library from tomato plants (Lycopersicon esculentum) with a cDNA probe encoding a subtilisin-like protease (PR-P69) that is induced at the transcriptional level following pathogen attack (Tornero, P., Conejero, V., and Vera, P. (1996) Proc. Natl. Acad. Sci. U. S. A. 93, 6332-6337) resulted in the isolation of a cluster of genomic clones that comprise a tandem of four different subtilisin-like protease genes (P69A, P69B, P69C, and P69D). Sequence analyses and comparison of the encoded proteins revealed that all are closely related (79 to 88% identity), suggesting that all are derived from a common ancestral gene. mRNA expression analysis as well as studies of transgenic plants transformed with promoter-␤-glucuronidase fusions for each of these genes revealed that the four genes exhibit differential transcriptional regulation and expression patterns. P69A and P69D are expressed constitutively, but with different expression profiles during development, whereas the P69B and P69C genes show expression following infection with Pseudomonas syringae and are also up-regulated by salicylic acid. We propose that these four P69-like proteases, as members of a complex gene family of plant subtilisin-like proteases, may be involved in a number of specific proteolytic events that occur in the plant during development and/or pathogenesis.Proteolysis is fundamental for the normal functioning of multicellular organisms and plays key roles in a variety of processes such as development, physiology, defense and stress responses, and adaptation to the changing environment. In plants, despite the importance of all these processes and involvement of different classes of proteinases (Refs. 1-5, and references therein), it still remains to be defined more precisely what components and molecular mechanisms are responsible for regulating specific aspects of protein degradation/processing. A major task for research will be to determine which pathway of proteolysis is responsible for the degradation of particular proteins.The serine proteases are one of the best characterized groups of proteolytic enzymes in higher organisms. They can be grouped in six clans, of which one of the largest is the subtilisin-like clan (EC 3.4.21.14) that includes over 200 different members. Despite this wealth of knowledge, very little is know about subtilisin-like proteases in plants. Recently, we and others have shown the existence of members of this clan in plants, including Arabidopsis (6), tomato (7, 8), melon (9), and Lilium plants (10). According to a recent classification (11), the subtilisin-like proteases from plants can be grouped within the Pyrolysin subfamily, which is highly related to the Kexin subfamily of proteases involved in the posttranslational processing of peptide hormones (12, 13). Comparative molecular, biochemical, and cellular studies indicate that the subgroup of plant subtilisin-like enzymes are characterized by the presence of a large polypeptide sequence insertion preceding the reactive Ser residue and/or lon...