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MicroRNAs (miRNAs) are recognized as essential transcriptional or post-transcriptional regulators, and play versatile roles in plants, functioning in processes such as growth, development and stress responses. Cassava (Manihot esculenta) is a major root crop widely grown as a staple food, animal feed, and also as an important source of bioethanol worldwide. Cold stress seriously affects cassava plants growth, development and yield. MiRNAs and their targets have been extensively studied in model plants, but a genomewide identification of miRNAs’ targets is still lacking in cassava. In this study, to identify the roles of miRNAs and their targets in response to cold stress, two degradome libraries were constructed using cold-treated and non-cold-treated cassava seedlings. Degradome data allowed us to identify a total of 151 non-redundant miRNA-target pairs from the degradome data. We reveal that approximately 42% of miRNA targets are conserved across plant species. However, 83 novel miRNA targets were identified in the two libraries, suggesting a specific role for these genes in response to cold stress. Gene ontology analyses showed that many target genes involved in “cellular process” and “metabolic process”. In addition, 12 miRNAs and 31 corresponding targets of them were further found to be involved in cold stress response. Particularly, miR159, miR164 and miR396 participated in cold stress response by up-regulating certain MYBs, NACs and GRFs genes that were involved in the regulation of downstream gene expression. Meantime, three miRNAs and five target genes were validated by quantitative real-time PCR (qRT-PCR). The work helps identify cold-responsive miRNA targets in cassava and increases the number of novel targets involved in cold stress response. Furthermore, the findings of this study might provide valuable reference and new insights for understanding the functions of miRNA in stress response in plants.

Genome-wide identification of miRNAs targets involved in cold response in cassava