Over-Expression of GmGIa-Regulated Soybean miR172a Confers Early Flowering in Transgenic Arabidopsis thaliana.

Tao Wang,Yong-Guang Li,Ming-Yang Sun,Wen-Bin Li,Xue-Song Wang

doi:10.3390/ijms17050645

Tao Wang, Yong-Guang Li + Show 3 more

Open Access

https://doi.org/10.3390/ijms17050645

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Abstract

Flowering is a pivotal event in the life cycle of plants. miR172 has been widely confirmed to play critical roles in flowering time control by regulating its target gene expression in Arabidopsis. However, the role of its counterpart in soybean remains largely unclear. In the present study, we found that the gma-miR172a was regulated by a GIGANTEA ortholog, GmGIa, in soybean through miRNA metabolism. The expression analysis revealed that gma-miR172a has a pattern of diurnal rhythm expression and its abundance increased rapidly as plants grew until the initiation of flowering phase in soybean. One target gene of gma-miR172a, Glyma03g33470, was predicted and verified using a modified RLM 5′-RACE (RNA ligase-mediated rapid amplification of 5′ cDNA ends) assay. Overexpression of gma-miR172a exhibited an early flowering phenotype and the expression of FT, AP1 and LFY were simultaneously increased in gma-miR172a-transgenic Arabidopsis plants, suggesting that the early flowering phenotype was associated with up-regulation of these genes. The overexpression of the gma-miR172a-resistant version of Glyma03g33470 weakened early flowering phenotype in the toe1 mutant of Arabidopsis. Taken together, our results suggested that gma-miR172a played an important role in GmGIa-mediated flowering by repressing Glyma03g33470, which in turn increased the expression of FT, AP1 and LFY to promote flowering in soybean.

Highlights

The timing of the switch from vegetative to reproductive growth is critical for the success of plant reproduction
Flowering time is regulated by coordinated interactions between various endogenous signals and environmental cues [1,2,3]
Genetic and molecular analyses had revealed that four major pathways regulate this transition: the endogenous factors include autonomous and the gibberellin pathways, while the photoperiod and vernalization pathways respond to environmental cues [4,5,6,7]

Summary

Introduction

The timing of the switch from vegetative to reproductive growth is critical for the success of plant reproduction. Genetic and molecular analyses had revealed that four major pathways regulate this transition: the endogenous factors include autonomous and the gibberellin pathways, while the photoperiod and vernalization pathways respond to environmental cues [4,5,6,7] These pathways are generally controlled by multiple genes and are influenced by the environment. Overexpression of miR156 reduces the level of target SPL genes and causes a late-flowering phenotype [12,13]. When miR159 was overexpressed, plants flowering time was delayed in SD condition with decreased levels of MYB33 and LFY in Arabidopsis [15]. In Arabidopsis, miR172 serves as a negative regulator of AP2 to specify floral organ identity and acts as a repressor of the AP2-like genes, the Target of EAT 1 (TOE1) and SCHLAFMU TZE (SMZ) to promote early flowering [19,20,21]. Toe mutant plants of Arabidopsis could restore its earlier flowering phenotype partially by the expression of Glyma03g33470

Identification and Analysis of gma-miR172a and Glyma03g33470 Sequences

Plant Materials and Growth Conditions

Real Time RT-PCR Analyses

RACE Mapping of miRNA Target Cleavage Sites

Gene Constructs and Generation of Transgenic Arabidopsis Plants

Flowering Time Measurements