Downregulation of miR156-Targeted PvSPL6 in Switchgrass Delays Flowering and Increases Biomass Yield.

Jinjun Cai,Wenwen Liu,Weiqian Li,Xinchang Zhang,Yangyang Bai,Gang Chen,Dongmei Ma,Lijuan Zhao,Chunxiang Fu,Yamei Wang

doi:10.3389/fpls.2022.834431

Abstract

MiR156/SQUAMOSA PROMOTER BINDING-LIKEs (SPLs) module is the key regulatory hub of juvenile-to-adult phase transition as a critical flowering regulator. In this study, a miR156-targeted PvSPL6 was identified and characterized in switchgrass (Panicum virgatum L.), a dual-purpose fodder and biofuel crop. Overexpression of PvSPL6 in switchgrass promoted flowering and reduced internode length, internode number, and plant height, whereas downregulation of PvSPL6 delayed flowering and increased internode length, internode number, and plant height. Protein subcellular localization analysis revealed that PvSPL6 localizes to both the plasma membrane and nucleus. We produced transgenic switchgrass plants that overexpressed a PvSPL6-GFP fusion gene, and callus were induced from inflorescences of selected PvSPL6-GFPOE transgenic lines. We found that the PvSPL6-GFP fusion protein accumulated mainly in the nucleus in callus and was present in both the plasma membrane and nucleus in regenerating callus. However, during subsequent development, the signal of the PvSPL6-GFP fusion protein was detected only in the nucleus in the roots and leaves of plantlets. In addition, PvSPL6 protein was rapidly transported from the nucleus to the plasma membrane after exogenous GA3 application, and returned from the plasma membrane to nucleus after treated with the GA3 inhibitor (paclobutrazol). Taken together, our results demonstrate that PvSPL6 is not only an important target that can be used to develop improved cultivars of forage and biofuel crops that show delayed flowering and high biomass yields, but also has the potential to regulate plant regeneration in response to GA3.

Highlights

Flowering is the key physiological juncture at which the plant transitions from vegetative to reproductive growth, and flowering at the optimum time is important for plant growth and reproductive success
In contrast to the extensive studies in Arabidopsis, little information is available on the flowering-related roles of SQUAMOSA PROMOTER BINDING-LIKE (SPL) in the Gramineae
Blastn searches against the switchgrass genome (P. virgatum v4.1, Phytozome) indicated that these four OG2 genes, PvSPL6 (Pavir.2KG430400), PvSPL7′ (Pavir.2NG503700), PvSPL8 (Pavir.2KG430000), and PvSPL17 (Pavir.2NG503500), were located on chromosome 2

Summary

Introduction

Flowering is the key physiological juncture at which the plant transitions from vegetative to reproductive growth, and flowering at the optimum time is important for plant growth and reproductive success. The regulatory mechanism of flowering time control in the model plant Arabidopsis thaliana has been studied extensively. It involves five major pathways: the photoperiodic, gibberellin, autonomous, vernalization, and age pathways. MiR156 and its target SQUAMOSA PROMOTER BINDING-LIKE (SPL) genes constitute the key regulatory hub of the age pathway (Fu et al, 2012; Teotia and Tang, 2015). FT is induced by the photoperiodic pathway and regulated by FUL and SOC1 under long-day conditions (Litt and Irish, 2003; Mathieu et al, 2007); and (3) SPL2/10/11, which have close homology to SPL9, can affect the flowering process by regulating FUL gene expression (Wang et al, 2009). PvSPL7 and 8 induce phase transition and flowering in grasses by directly upregulating SEPALLATA3 (SEP3) and MADS32 (Gou et al, 2019)

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