miR393, which is encoded by MIR393a and MIR393b in Arabidopsis, post-transcriptionally regulates mRNAs for the F-box auxin receptors TIR1 (Transport Inhibitor Response Protein 1), AFB1 (Auxin Signaling F-box Protein 1), AFB2 and AFB3. However, biological functions of the miR393-TIR1/AFBs module in auxin response and plant development is not fully understood. In the study herein, we demonstrate that miR393 accumulated in response to exogenous IAA treatment, and its induction was due to enhanced MIR393b transcription but not MIR393a. Overexpression of a miR393-resistant form of TIR1 (mTIR1) enhanced auxin sensitivity and led to pleiotropic effects on plant development including inhibition of primary root growth, overproduction of lateral roots, altered leave phenotype and delayed flowering. Furthermore, miR393 level was increased in 35S:mTIR1 plant, suggesting that TIR1 promoted the expression of miR393 by a feedback loop. The interaction between miR393 and its target indicates a fine adjustment to the roles of the miR393-TIR1 module, which is required for auxin responses in plant development.

miR396 targets seven GROWTH-REGULATING FACTOR (GRF) genes and the BASIC HELIX-LOOP-HELIX (bHLH) TRANSCRIPTION FACTOR 74 gene (bHLH74) in Arabidopsis. Previous research revealed that the miR396 target module regulates cell proliferation and plays a critical role in leaf development. However, no additional biological functions of miR396 have been investigated in detail. In this study, T-DNA insertion mutants and transgenic plants with altered levels of miR396 or its target genes were used to characterize the regulatory role of miR396 in root development. We found that AtMIR396a was the predominant source for miR396 accumulation in the roots of seedlings, and that the mir396a-1 mutant had longer roots than wild-type seedlings. Overexpression of AtMIR396a decreased the transcript levels of target genes such as GRF genes and bHLH74, and resulted in a shorter root phenotype. Furthermore, the bhlh74-1 mutant had shorter roots, whereas overexpression of an miR396-resistant form of bHLH74 (mbHLH74) had an enhanced root growth phenotype. Moreover, MIR396a regulated root growth by affecting the elongation zone. Taken together, these data indicate that miR396a-mediated bHLH74 repression helps regulate root growth in Arabidopsis seedlings.