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Abstract Diketopiperazines (DKPs) are chemically and functionally diverse cyclic dipeptides associated primarily with microbes. Few DKPs have been reported from plants and animals; the best characterized is cyclo(His-Pro), found in the mammalian central nervous system, where it arises from the proteolytic cleavage of a thyrotropin-releasing tripeptide hormone. Herein, we report the identification of cyclo(His-Pro) in Arabidopsis (Arabidopsis thaliana), where its levels increase upon abiotic stress conditions, including high salt, heat, and cold. To screen for potential protein targets, we used isothermal shift assays, which examine changes in protein-melting stability upon ligand binding. Among the identified proteins, we focused on the glycolytic enzyme, cytosolic glyceraldehyde-3-phosphate dehydrogenase (GAPC1). Binding between the GAPC1 protein and cyclo(His-Pro) was validated using nano-differential scanning fluorimetry and microscale thermophoresis, and we could further demonstrate that cyclo(His-Pro) inhibits GAPC1 activity with an IC50 of ∼200 μm. This inhibition was conserved in human GAPDH. Inhibition of glyceraldehyde-3-phosphate dehydrogenase activity has previously been reported to reroute carbon from glycolysis toward the pentose phosphate pathway. Accordingly, cyclo(His-Pro) supplementation augmented NADPH levels, increasing the NADPH/NADP+ ratio. Phenotypic screening revealed that plants supplemented with cyclo(His-Pro) were more tolerant to high-salt stress, as manifested by higher biomass, which we show is dependent on GAPC1/2. Our work reports the identification and functional characterization of cyclo(His-Pro) as a modulator of glyceraldehyde-3-phosphate dehydrogenase in plants. 

Summary Drought stress substantially impacts crop physiology resulting in alteration of growth and productivity. Understanding the genetic and molecular crosstalk between stress responses and agronomically important traits such as fibre yield is particularly complicated in the allopolyploid species, upland cotton (Gossypium hirsutum), due to reduced sequence variability between A and D subgenomes. To better understand how drought stress impacts yield, the transcriptomes of 22 genetically and phenotypically diverse upland cotton accessions grown under well‐watered and water‐limited conditions in the Arizona low desert were sequenced. Gene co‐expression analyses were performed, uncovering a group of stress response genes, in particular transcription factors GhDREB2A‐A and GhHSFA6B‐D, associated with improved yield under water‐limited conditions in an ABA‐independent manner. DNA affinity purification sequencing (DAP‐seq), as well as public cistrome data from Arabidopsis, were used to identify targets of these two TFs. Among these targets were two lint yield‐associated genes previously identified through genome‐wide association studies (GWAS)‐based approaches,GhABP‐DandGhIPS1‐A. Biochemical and phylogenetic approaches were used to determine thatGhIPS1‐Ais positively regulated by GhHSFA6B‐D, and that this regulatory mechanism is specific toGossypiumspp. containing the A (old world) genome. Finally, an SNP was identified within the GhHSFA6B‐D binding site inGhIPS1‐Athat is positively associated with yield under water‐limiting conditions. These data lay out a regulatory connection between abiotic stress and fibre yield in cotton that appears conserved in other systems such as Arabidopsis.