MicroRNAs are small RNAs, 20–22 nt long, the main role of which is to downregulate gene expression at the level of mRNAs. MiRNAs are fundamental regulators of plant growth and development in response to internal signals as well as in response to abiotic and biotic factors. Therefore, the deficiency or excess of individual miRNAs is detrimental to particular aspects of a plant’s life. In consequence, the miRNA levels must be appropriately adjusted. To obtain proper expression of each miRNA, their biogenesis is controlled at multiple regulatory layers. Here, we addressed processes discovered to influence miRNA steady-state levels, such as MIR transcription, co-transcriptional pri-miRNA processing (including splicing, polyadenylation, microprocessor assembly and activity) and miRNA-encoded peptides synthesis. MiRNA stability, RISC formation and miRNA export out of the nucleus and out of the plant cell also define the levels of miRNAs in various plant tissues. Moreover, we show the evolutionary conservation of miRNA biogenesis core proteins across the plant kingdom.

HYL1 (DRB1) is a double-stranded RNA binding protein involved in miRNA processing in plants. It is a core component of the Microprocessor complex and enhances the efficiency and precision of miRNA processing by the DCL1 protein. In this work, we report a novel function of the HYL1 protein in the transcription of MIR genes. HYL1 colocalizes with RNA Polymerase II and affects its distribution along MIR genes. Moreover, proteomic experiments revealed that the HYL1 protein interacts with many transcription factors. Finally, we show that the action of HYL1 is not limited to MIR genes and impacts the expression of many other genes, a majority of which are involved in the plastid organization. These discoveries indicate HYL1 as an additional player in gene regulation at the transcriptional level, independent of its role in miRNA biogenesis.

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