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In this study, we investigated the regulatory function of miR396 in the phase transition in Arabidopsis thaliana. Using AtMIR396a/b knockout mutants generated through clustered regularly interspaced short palindromic repeats/CRISPR-associated protein 9 (CRISPR/Cas9)-directed genome editing, we showed that miR396 negatively regulates the leaf size and vegetative phase transition, and the first leaf with abaxial trichomes appeared earlier in the mir396ab double mutant than in the wild type (WT) and was significantly delayed in miR396 overexpression lines. Moreover, mir396ab exhibited early flowering, whereas 35S:MIR396a/b and cib4-1 delayed flowering, and the flowering time was negatively correlated with FT gene expression. Furthermore, in arp6 and pie1 mutants, which are deficient in the ATP-dependent chromatin remodeling complex (SWR1-C), miR396 expression was significantly repressed. Compared with the WT, reduced H2A.Z deposit and stronger relative nucleosome occupancy in the promoter region of MIR396a was found in the arp6 mutant, indicating that SWR1-C contributes to the transcriptional activation of MIR396a via nucleosome dynamics. In addition, miR396 displayed specific spatio-temporal expression patterns in the leaf, which was altered in arp6 and pie1 , and therefore affected the transcript levels of CIB4 and FT in these mutants. We propose that miR396 is not only a marker of cell differentiation, but also an age signal for leaf development and phase change. Meanwhile, SWR1-C-mediated epigenetic regulation contributes to the age-dependent enhancement of miR396 expression and differential miR396 accumulation among leaves. [ABSTRACT FROM AUTHOR]

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. [ABSTRACT FROM PUBLISHER]

Among the three primary auxin-induced gene families, Auxin/Indole-3-Acetic Acid (Aux/IAA), Gretchen Hagen3 (GH3) and SMALL AUXIN UP RNA (SAUR), the function of SAUR genes remains unclear. Arabidopsis SAUR genes have been phylogenetically classified into three clades. Recent work has suggested that SAUR19 (clade II) and SAUR63 (clade I) promote cell expansion through the modulation of auxin transport. Herein, we present our work on SAUR41, a clade III SAUR gene with a distinctive expression pattern in root meristems. SAUR41 was normally expressed in the quiescent center and cortex/endodermis initials; upon auxin stimulation, the expression was provoked in the endodermal layer. During lateral root development, SAUR41 was expressed in prospective stem cell niches of lateral root primordia and in expanding endodermal cells surrounding the primordia. SAUR41–EGFP (enhanced green fluorescent protein) fusion proteins localized to the cytoplasm. Overexpression of SAUR41 from the Cauliflower mosaic virus 35S promoter led to pleiotropic auxin-related phenotypes, including long hypocotyls, increased vegetative biomass and lateral root development, expanded petals and twisted inflorescence stems. Ectopic SAUR41 proteins were able to promote auxin transport in hypocotyls. Tissue-specific expression of SAUR41 from the PIN1, WOX5, PLT2 and ACR4 promoters induced the formation of new auxin accumulation/signaling peaks above the quiescent centers, whereas tissue-specific expression of SAUR41 from the PIN2 and PLT2 promoters enhanced root gravitropic growth. Cells in the root stem cell niches of these transgenic seedlings were differentially enlarged. The distinctive expression pattern of the SAUR41 gene and the explicit function of SAUR41 proteins implied that further investigations on the loss-of-function phenotypes of this gene in root development and environmental responses are of great interest. [ABSTRACT FROM PUBLISHER]