Genome-Wide Identification of miRNAs and Pollen-Associated miRNA-Target Relationships in Liriodendron chinense

MicroRNAs (miRNAs) are key post-transcriptional regulators of gene expression and play important roles in plant development and reproduction. Liriodendron chinense, a representative woody species of Magnoliaceae, produces a large quantity of pollen but exhibits low natural seed set. Despite numerous studies on factors influencing its low seed production, the molecular mechanisms underlying this reproductive limitation remain poorly understood. Here, we performed small RNA sequencing and transcriptome analyses across five tissues of L. chinense, including leaf, calyx, petal, pistil, and pollen. Genome-wide identification yielded 688 miRNAs, comprising both conserved and species-specific members. Expression-based clustering revealed that miRNAs are organized into distinct, tissue-associated modules rather than being uniformly expressed across organs. Among these, a pollen-enriched miRNA module was clearly separated from those associated with leaves and other floral tissues. By integrating sRNA-seq and RNA-seq data, we identified miRNA-target pairs displaying anticorrelated expression patterns, providing expression-level support for miRNA mediated regulation. In pollen, two complementary regulatory modes were observed: low-abundance miRNAs associated with highly expressed target genes, and highly expressed miRNAs associated with repressed targets. The predicted targets of pollen-associated miRNAs were enriched in biological processes central to pollen development, including signal transduction, polarity establishment, vesicle trafficking, and cell wall biogenesis. Overall, this study provides a comprehensive, tissue-resolved view of miRNA expression and pollen-associated miRNA-target relationships in L. chinense, offering candidate regulatory modules for future functional studies of pollen development.