Epigenetic regulation of microRNA395 expression under sulfate deprivation in Arabidopsis thaliana

MicroRNAs fine-tune target gene expression at post-transcriptional level in organ-, cell-, developmental stage- and stress-specific manner. MicroRNA395 (miR395) targets transcripts encoding two different classes of proteins, i.e., a sulfate transporter (AST68) and ATP sulfurylases (APS1, APS3 and APS4) in Arabidopsis. Previous studies have shown that sulfate deprivation (S deprivation) induces miR395 expression in Arabidopsis. This induced expression is transcriptionally controlled by SULFUR LIMITATION 1, a transcription factor. However, the epigenetic processes that are associated with miR395 induction under S deprivation are unknown. Histone methylation and histone variants play an essential role in regulating gene expression by altering the chromatin state. In this study, the role of H3K4me3 (ATX1), H3K4me2 (ATX2) and H3K27me3 (SWN3) as well as the role of H2A.Z variant (HTA9 and HTA11) in regulating miR395 induction was assessed in mutants that are defective in depositing these epigenetic marks.The analysis of all six pri-MIR395 transcripts (miR395a, b, c, d, e and f) revealed all of these loci are strongly up-regulated in shoots, however in roots, four of the six loci exhibited up-regulation under S deprivation. However, in mutants lacking H3K4me3 and H3K4me2 and their combination (atx1-2, atx2-1 and atx1-2/atx2-1), the expression levels were significantly different; 5 and 4 loci in shoots and roots, respectively, were more strongly up-regulated in mutants compared to Col-0. Similarly, in mutants lacking H3K27me3 (swn3), 3 (shoots) and 4 (roots) loci in shoots and roots, respectively, showed more up-regulation compared to Col-0. Compared to the Col-0 responses, the pri-MIR395 levels also showed differences both in shoots and roots of hta single mutants, and the differences were more pronounced in hta9-1/hta11-2 double mutants.Overall, some of the results indicated that these epigenetic marks are important for regulating some of the miR395 loci while some other loci did not agree with the expectations but showed contradictory results. It is possible that the ATX1, ATX2, SWN3, HTA9 and HTA11 genes that are analyzed in the present study is only targeting sub-set of miR395 loci in Arabidopsis