Identification and experimental validation of a circular RNA associated with demethylase genes in maize

In the context of climate change and the need to reduce pesticide and agrochemical use, the development of sustainable crops is becoming increasingly important. Epigenetic mechanisms, such as histone modifications, allow plants to adapt to stress without altering their DNA sequence. Histone demethylases are of particular interest as enzymes that activate defense genes. Concurrently, circular RNAs (circRNAs) have emerged as a new class of regulatory molecules capable of modulating gene expression. The aim of this study was to identify and validate circRNAs derived from histone demethylase genes in maize (Zea mays L.) during early developmental stages. The research was conducted on 10-day-old seedlings. Based on annotated genes and the PlantcircBase 7.0 database, six predicted circRNAs were selected. They were validated using PCR with divergent primers and subsequent sequencing. The formation of only one circular RNA out of the six predicted was experimentally confirmed – from the locus of the demethylase gene Zm00001d051961. It has been established that this circRNA is formed through backsplicing between the 2nd and the 7th exons, is 2016 nucleotides in size, and exhibits tissue-specific expression, predominantly in the leaves. This is the first experimental proof of a circRNA originating from a histone demethylase gene locus in maize. The discovered tissue-specific expression suggests that this circRNA may be involved in the fine-tuning of the demethylase's activity, potentially influencing the epigenetic status of chromatin. This discovery enhances the understanding of plant epigenetic plasticity and lays the groundwork for developing new breeding strategies aimed at enhancing crop stress resilience.