MicroRNA390-Directed TAS3 Cleavage Leads to the Production of tasiRNA-ARF3/4 During Somatic Embryogenesis in Dimocarpus longan Lour.

Yuling Lin,Zhongxiong Lai,Ruilian Lai,Weihua Liu,Xu Xuhan,Yukun Chen,Lixia Lin,Zihao Zhang

doi:10.3389/fpls.2015.01119

Yuling Lin, Zhongxiong Lai + Show 6 more

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https://doi.org/10.3389/fpls.2015.01119

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Abstract

Trans-acting short-interfering RNAs (tasiRNAs) originate from TAS3 families through microRNA (miRNA) 390-guided cleavage of primary transcripts and target auxin response factors (ARF3/-4), which are involved in the normal development of lateral roots and flowers in plants. However, their roles in embryo development are still unclear. Here, the pathway miR390-TAS3-ARF3/-4 was identified systematically for the first time during somatic embryo development in Dimocarpus longan. We identified the miR390 primary transcript and promoter. The promoter contained cis-acting elements responsive to stimuli such as light, salicylic acid, anaerobic induction, fungal elicitor, circadian control, and heat stress. The longan TAS3 transcript, containing two miR390-binding sites, was isolated; the miR390- guided cleavage site located near the 3′ end of the TAS3 transcript was verified. Eight TAS3-tasiRNAs with the 21-nucleotides phase were found among longan small RNA data, further confirming that miR390-directed TAS3 cleavage leads to the production of tasiRNA in longan. Among them, TAS3_5′D5+ and 5′D6+ tasiRNAs were highly abundant, and verified to target ARF3 and -4, implying that miR390-guided TAS3 cleavage with 21-nucleotides phase leading to the production of tasiRNA-ARF is conserved in plants. Pri-miR390 was highly expressed in friable-embryogenic callus (EC), and less expressed in incomplete compact pro-embryogenic cultures, while miR390 showed its lowest expression in EC and highest expression in torpedo-shaped embryos (TEs). DlTAS3 and DlARF4 both exhibited their lowest expressions in EC, and reached their peaks in the globular embryos stage, which were mainly inversely proportional to the expression of miR390, especially at the globular embryos to cotyledonary embryos (CEs) stages. While DlARF3 showed little variation from the EC to TEs stages, and exhibited its lowest expression in the CEs stage. There was a general lack of correlation between the expressions of DlARF3 and miR390. In addition, pri-miR390, DlTAS3, DlARF3 and -4 were up-regulated by 2,4-D in a concentration-dependent manner. They were also preferentially expressed in roots, pulp, and seeds of ‘Sijimi’ longan, implying their extended roles in the development of longan roots and fruit. This study provided insights into a possible role of miR390-tasiRNAs-ARF in plant somatic embryo development.

Highlights

MicroRNAs and tasiRNAs are distinct classes of small RNAs, which control many aspects of development in plants by guiding silencing of target RNAs via cleavage or repression mechanisms (Chapman and Carrington, 2007)
To determine miR390s structure and the length of the gene in the D. longan genome, the full-length complementary DNA (cDNA) sequence of the primary miR390 transcript was obtained by reverse transcription-polymerase chain reaction (RT-PCR) and 5 -RACE
TasiRNAs originate from TAS3 families through miR390-guided initiation-cleavage of primary transcripts and target ARF2/-3/-4, which are involved in the normal development of lateral roots (Marin et al, 2010;Yoon et al, 2010) and flowers in plants (Matsui et al, 2014); their roles in embryo development are still unclear

Summary

Introduction

MicroRNAs and tasiRNAs are distinct classes of small RNAs, which control many aspects of development in plants by guiding silencing of target RNAs via cleavage or repression mechanisms (Chapman and Carrington, 2007). MiRNAs arise from endogenous transcripts that can form local fold-back structures, whereas tasiRNAs are generated by primary TAS transcripts that are initially targeted and sliced by miR173 (TAS1 and TAS2), miR828 (TAS4) or miR390 (TAS3) (Pullman et al, 2003; Wilson et al, 2005; Axtell et al, 2006; Kikuchi et al, 2006; Montgomery et al, 2008b). TAS3 tasiRNAs target ARF3 and -4, which are involved in developmental timing (Cho et al, 2012) and the normal development of aerial lateral organs, such as lateral roots (Marin et al, 2010; Yoon et al, 2010) and flowers (Matsui et al, 2014)

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