Dynamic and Functional miRNA‐Mediated Gene Regulations in Response to Recurrent Environmental Challenges During Biological Invasions

Biological invasions offer a valuable 'natural experiment' to investigate survival mechanisms, as invaders successfully endure substantial environmental changes during their geographical spread and settlement. Phenotypic plasticity enhances fitness by enabling rapid responses without requiring new genetic variations. Among numerous mechanisms involved in phenotypic plasticity, microRNAs (miRNAs) and their regulatory networks are believed to enable rapid responses by fine-tuning gene expression, though their roles remain poorly understood. By integrating miRNAomic and transcriptomic analyses in the model invasive ascidian Ciona robusta, we simulated recurring salinity stresses encountered during invasions to investigate the molecular mechanisms of miRNA-mediated gene regulation in response to recurrent environmental challenges. Multiple analyses demonstrated that miRNAs exhibited rapid, dynamic and reversible responses to recurrent stresses, displaying duration-dependent and stage-specific profiles. The upregulation of genes in the miRNA biogenesis process, rather than the decay pathway, primarily accounted for the increased expression abundance of miRNAs. Responsive miRNAs regulated target genes through an intricate regulatory network, demonstrated by both up and downregulatory relationships and diverse binding sites. Interestingly, miRNAs and their target genes exhibited a 'stress memory' effect, where miRNAs 'remembered' previous challenges and further mediated the enhanced response of target genes at later stresses. Functionally, miRNA-mediated salinity coping strategies and associated genes exhibited temporal variations depending on challenge duration and stage, and these strategies primarily involved the modulation and alternation of free amino acid metabolism and ion transport to maintain osmotic homeostasis. These findings highlight the importance of miRNA-mediated regulatory networks in shaping short-term phenotypic plasticity in response to environmental changes.