Abstract
MicroRNAs (miRNAs) are a class of non-coding RNAs that play important roles in plant development and abiotic stresses. To date, studies have mainly focused on the roles of individual miRNAs, however, a few have addressed the interactions among multiple miRNAs. In this study, we investigated the interplay and regulatory circuit between miR160 and miR165/166 and its effect on leaf development and drought tolerance in Arabidopsis using Short Tandem Target Mimic (STTM). By crossing STTM160 Arabidopsis with STTM165/166, we successfully generated a double mutant of miR160 and miR165/166. The double mutant plants exhibited a series of compromised phenotypes in leaf development and drought tolerance in comparison to phenotypic alterations in the single STTM lines. RNA-seq and qRT-PCR analyses suggested that the expression levels of auxin and ABA signaling genes in the STTM-directed double mutant were compromised compared to the two single mutants. Our results also suggested that miR160-directed regulation of auxin response factors (ARFs) contribute to leaf development via auxin signaling genes, whereas miR165/166- mediated HD-ZIP IIIs regulation confers drought tolerance through ABA signaling. Our studies further indicated that ARFs and HD-ZIP IIIs may play opposite roles in the regulation of leaf development and drought tolerance that can be further applied to other crops for agronomic traits improvement.
Highlights
MicroRNAs are small, endogenous, non-coding RNAs that function in gene regulation by mRNAs cleavage or translational repression in plants[1]
The expression of miR160 was slightly reduced in STTM165/166 single mutant and that of miR165/166 slightly increased in STTM160 single mutant compared to wild type (Fig. 3A,C)
Our results revealed that the expression of TRYPTOPHAN AMINOTRANSFERASE OF ARABIDOPSIS 1 (TAA1) was significantly lower in double mutant whereas that of YUCCA 1 (YUC1) were higher in double mutant in comparison to STTM160 (Fig. 5B)
Summary
MicroRNAs (miRNA) are small, endogenous, non-coding RNAs that function in gene regulation by mRNAs cleavage or translational repression in plants[1]. In Arabidopsis, STTM165/166 plants displayed multiple morphological phenotypes including twisted inflorescence stems, altered cauline branches, dark purple leaves, reduced fertility and delayed flowering These plants have increased indole acetic acid (IAA) contents and decreased IAA sensitivity, suggesting the possible roles of miR165/166 in auxin biosynthesis and signaling[10]. The STTM166 plants showed high drought tolerance due to reduced stomatal conductance and transpiration rates[6] Both miR160 and miR165/166 are involved in the modulation of leaf primordium initiation, leaf polarity establishment and abiotic stress responses. STTM160 and STTM165/166 Arabidopsis transgenic plants and their double mutants were generated and used to decipher their functional interactions and their specific roles in leaf development associated with auxin signaling and the ABA signaling-associated abiotic stresses. Our findings revealed distinct miRNA-regulatory networks between STTM160 and STTM165/166 and the interactions of these two miRNA-guided gene networks in the double mutant
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