Heat-responsive microRNAs regulate the transcription factors and heat shock proteins in modulating thermo-stability of starch biosynthesis enzymes in wheat (Triticum aestivum L.) under the heat stress

Heat-responsive miRNAs regulate the expression of the transcription factors (HSFs) and majority of the heat stress-associated genes (HSPs). Here we report identification of few heats responsive miRNAs in wheat (Triticum aestivum L.)through de novo sequencing on Illumina Hiseq 2000. Validation of identified miRNAs in endospermic tissues of thermotolerant (HD2985) and thermosusceptible (NIAW-34-34) wheat cultivars using real time PCR showed up-regulation of 4 micro RNAs (tae-miR156, tae-miR167, tae-miR395b and tae-miR398) and down-regulation of 6 micro RNAs (tae-miR159a, tae-miR159b, tae-miR160, taemiR171a, tae-miR319, and tae-miR1117) in response to the heat stress. Target analysis of identified miRNAs showed HSF3, HSFA4a, HSP17, HSP70 and superoxide dismutase (SOD) as most probable target genes. Expression profiling of identified target genes under heat stress (42°C, 2 h) showed 2.34, 1.33 fold (HSF3), 2.45, 1.44 fold (HSFA4a), 3.9, 1.9 fold (HSP17), 5.6, 2.4 fold (HSP70), 1.9, 1.2 fold (SOD) and 2.7, 1.6 fold (catalase) increase in the expression in HD2985 and NIAW-34-34 cultivars of wheat compared to control. A defragmented, small and pleated starch granule structure was observed in sample with low expression of target genes (NIAW-34-34) compared to intact, robust and globular starch granules in samples with high expression of target genes (HD2985). Transcript profiling and activity assay of soluble starch synthase (SSS) showed less transcript accumulation and activity in heat shock treated samples compared to control sample in both the cultivars.