Multiple factors control primary seed dormancy established during seed maturation and secondary seed dormancy initiated when a non-dormant imbibed seed is exposed to adverse conditions. A key player in the control of primary and secondary dormancy in Arabidopsis thaliana is the Delay of Germination 1 (DOG1) gene, the expression of which is extensively regulated at the transcriptional and co-transcriptional levels. Despite its importance, the influence of post-transcriptional messenger RNA (mRNA) processing and mRNA storage of DOG1 on the determination of dormancy depth remains elusive. Here, we show that the UBA2A protein, a member of the heterogeneous nuclear ribonucleoprotein (hnRNP) family, negatively regulates primary and secondary seed dormancy through the regulation of the DOG1 gene expression at the post-transcriptional level. uba2a mutants show higher levels of the DOG1 mRNA. Surprisingly, DOG1 gene transcription is not affected, as demonstrated by single-molecule fluorescent in situ hybridization, chromatin-attached mRNA analysis and Pol II chromatin immunoprecipitation (ChIP). Instead, our results show that the UBA2A protein decreases the stability of both chromatin-bound and cytoplasmic DOG1 mRNA pools, and results in higher chromatin retention of DOG1 mRNA in the uba2a mutant. Our study highlights chromatin retention and mRNA stability as important features of DOG1 gene expression regulation with a profound impact on dormancy establishment and shows that UBA2A protein, like its human homolog hnRNPAB, is most likely implicated in mRNA transport in the cell.

Inositol pyrophosphates (PP-InsPs) are important signaling molecules that regulate diverse cellular processes in eukaryotes, including energy homeostasis, phosphate (P_i) signaling, and phytohormone perception. Yet, in plants, the enzymes responsible for their turnover remain largely unknown. Using a non-hydrolysable PP-InsP analog in a pull-down approach, we identified a family of Arabidopsis NUDIX-type hydrolases (NUDTs) that group into two closely related subclades. Through in vitro assays, heterologous expression systems, and higher order gene-edited mutants, we explored the substrate specificities and physiological roles of these hydrolases. Using a combination of strong anion exchange high-performance liquid chromatography (SAX-HPLC), polyacrylamide gel electrophoresis (PAGE), and capillary electrophoresis electrospray ionization mass spectrometry (CE-ESI-MS), we found that their PP-InsP pyrophosphatase activity is enantiomer selective and Mg²⁺ dependent. Specifically, Subclade I NUDTs preferentially hydrolyze 4-InsP₇, while Subclade II NUDTs target 3-InsP₇, with minor activity against other PP-InsPs, including 5-InsP₇. In higher order mutants of Subclade II NUDTs, we observed defects in both P_i and iron homeostasis, accompanied by increased levels of 1/3-InsP₇ and 5-InsP₇, with a markedly larger increase in 1/3-InsP₇. Ectopic expression of NUDTs from both subclades induced local Pi starvation responses (PSRs), while RNA-seq analysis comparing wild-type (WT) and Subclade II nudt12/13/16 loss-of-function plants indicates additional PSR-independent roles, potentially involving 1/3-InsP₇ in the regulation of plant defense. Consistently, nudt12/13/16 mutants displayed enhanced resistance to Pseudomonas syringae infection, indicating a role in bacterial pathogen susceptibility. Expanding beyond Subclade II NUDTs, we demonstrated susceptibility of the 3PP-position of PP-InsPs to enzymatic activities unrelated to NUDTs, and found that such activities are conserved across plants and humans. Additionally, we observed that NUDT effectors from pathogenic ascomycete fungi exhibit a substrate specificity similar to Subclade I NUDTs. Collectively, our findings reveal new roles for NUDTs in PP-InsP signaling, plant nutrient and immune responses, and highlight a cross-kingdom conservation of PP-InsP-metabolizing enzymes.

Symbiotic nitrogen fixation in legumes requires the exquisite regulation of the environment within the infected region of the nodule. The microaerobic environment critical for nitrogenase activity is maintained through the physical oxygen diffusion barrier of the cortex and locally the oxygen-binding protein leghemoglobin (Lb). Leghemoglobin binds and releases oxygen with heme moiety to maintain oxygen gradients inside the infected cell (IC) during nitrogen fixation. Heme binds to diverse proteins and plays critical roles in different redox reactions. However, the role and regulation of host-controlled heme production during symbiotic nitrogen fixation are not clear. Here, we identified coproporphyrinogen III oxidase plastid related 1 (CPOP1) as a key regulator of symbiotic heme biosynthesis in Lotus japonicus. CPOP1 is specifically highly expressed in nitrogen-fixing nodules, and knocking out CPOP1 alone causes leaf etiolation and dwarfism which could be recovered by the exogenous application of nitrogen source, indicating nitrogen fixation defect. The IC-specific expression of CPOP1 was directed by the −881 to −740 bp promoter region. The cpop1 mutant shows significantly increased nodule oxygen level and decreased nitrogen fixation activity compared to the wild-type. Intriguingly, bacteria proliferation is inhibited due to the down-regulation of cell division-related gene expression upon CPOP1 knockout. Our data showed that CPOP1 is essential for the microaerobic environment control of ICs and the activation of rhizobial nitrogenase required for symbiotic nitrogen fixation, through host-regulated nodule heme synthesis.

The pollen exine serves as a protective barrier and signaling interface essential for male fertility in flowering plants. Its precise patterning depends on coordinated interactions between microspores and tapetal cells. While the CLAVATA3/EMBRYO SURROUNDING REGION-related 19 (CLE19) peptide has been identified as a microspore-derived “brake” that restricts tapetal activity to maintain exine developmental homeostasis, how CLE19 integrates with hormonal signaling pathways remains poorly understood. Here, we demonstrate that CLE19 attenuates brassinosteroid (BR) signaling output by engaging a defined BSL–BIN2–BES1 signaling cascade. Through quantitative phosphoproteomic analysis, we identified that CLE19 affects the phosphorylation of multiple BR signaling components, including BSL-type phosphatases BSL1/2/3, the GSK3-like kinase BIN2, and the transcription factor BES1. We show that CLE19 is perceived by its receptor PXL1, which directly interacts with BSL-type phosphatases to activate the GSK3-like kinase BIN2, leading to phosphorylation of BES1 at serine residues S219 and S223. Functional analyses using phospho-dead and phospho-mimic BES1 variants confirm that CLE19-dependent phosphorylation controls BES1 nuclear export and degradation, ultimately suppressing BR-responsive transcriptional outputs required for pollen exine patterning. Together, our findings define a peptide–hormone signaling axis that regulates transcription factor activity through post-translational regulation, providing mechanistic insight into how developmental robustness is maintained via intercellular signal integration in plant reproduction.