Circular RNAs (circRNAs) are noncoding RNAs formed by back-splicing, characterized by covalently closed-loop structures with enhanced stability. Although growing evidence highlights their regulatory roles in plants, the precise biological functions of circRNAs remain largely unclear. In this study, we identified Arabidopsis thaliana circP5CS1, a circRNA derived from the delta-1-pyrroline-5-carboxylate synthase (P5CS1) gene through back-splicing of exons 11 to 13. The junction between exons 11-13 generates a de novo miR167 binding site, validated by sequence alignment and RNA pull-down assays, which indicate that circP5CS1 preferentially binds miR167a/b. circP5CS1 acts as a miR167 sponge to upregulate auxin response factors 6 (ARF6) and ARF8, fine-tuning salicylic acid (SA)- and jasmonic acid (JA)-mediated immunity during Pseudomonas syringae pv. tomato (Pst) DC3000 infection. In addition, circP5CS1 suppresses its host gene P5CS1, a rate-limiting enzyme for proline biosynthesis, through both splicing competition and post-transcriptional mechanisms, thereby linking circRNA biogenesis with gene expression control. Our results suggest that circP5CS1 directly modulates plant immunity via both hormone and proline pathways. circP5CS1 expression was induced by both Pst DC3000 and AvrRpt2 infections. Consistent with this, resistance to these pathogens was compromised in circP5CS1-OE, p5cs1, and miR167-STTM plants, whereas circP5CS1-RNAi, P5CS1-1-OE, and miR167-OE showed enhanced resistance. Proline levels increased in P5CS1-1-OE and circP5CS1-RNAi plants but decreased in circP5CS1-OE and p5cs1 mutants, supporting a link between circP5CS1 activity and proline metabolism. Our findings suggest that circP5CS1 may be an immuno-hub integrating hormone signaling and proline homeostasis through miRNA sponging and host gene suppression, revealing its dual regulatory role in coordinating RNA-mediated regulation with plant immune response.
Wang et al. (Fri,) studied this question.