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. 2022 Mar 7;13(3):469.
doi: 10.3390/genes13030469.

Effect of the PmARF6 Gene from Masson Pine (Pinus massoniana) on the Development of Arabidopsis

Youju Ye  1   2 , Xin Han  1 , Hao Rong  1 , Renjuan Qian  1   2 , Jian Zheng  2 , Zhouxian Ni  1 , Li'an Xu  1
Affiliations

Effect of the PmARF6 Gene from Masson Pine (Pinus massoniana) on the Development of Arabidopsis

Youju Ye et al. Genes (Basel). .

Abstract

Masson pine (Pinus massoniana) is a core industrial tree species that is used for afforestation in southern China. Previous studies have shown that Auxin Response Factors (ARFs) are involved in the growth and development of various species, but the function of ARFs in Masson pine is unclear. In this research, we cloned and identified Masson pine ARF6 cDNA (PmARF6). The results showed that PmARF6 encodes a protein of 681 amino acids that is highly expressed in female flowers. Subcellular analysis showed that the PmARF6 protein occurred predominantly in the nucleus and cytomembrane of Masson pine cells. Compared with wild-type (WT) Arabidopsis, transgenic Arabidopsis plants overexpressing PmARF6 had fewer rosette leaves, and their flower development was slower. These results suggest that overexpression of PmARF6 may inhibit the flower and leaf development of Masson pine and provide new insights into the underlying developmental mechanism.

Keywords: Auxin Response Factor (ARF); Pinus massoniana; PmARF6; transgenic plant.

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Conflict of interest statement

The authors declare no conflict of interest.

Figures

Figure 1
Figure 1
Sequence analysis of the PmARF6 gene. (A) The gene structure of PmARF6. The 5′-UTR and 3′UTR are represented by the blue lines. The yellow line represents the 2043 bp ORF. Different cylinder-shaped colors represent different domains. The rectangle represents the Gln-rich region. (B) Sequence alignment of the ARF proteins. GenBank accession numbers: PsiARF6 (Picea sitchensis Psi012882), GbARF6 (Ginkgo biloba, CBA120031), AtARF6 (Arabidopsis thaliana, AT1G30330.1), PtrARF6 (Populus trichocarpa, Potri.001G358500.1), AtARF8 (Arabidopsis thaliana, AT5G37020.1), PtaARF2 (Pinus taeda, Pta011597), PaARF2 (Picea abies, MA_10432349g0010), and PaARF4 (Picea abies, MA_10431460g0020). (C) Phylogenetic tree of ARF6 proteins constructed with the neighbor-joining method in MEGA X. The G1, G2, G3 and G4 was represented by the light blue rectangle, red, dark blue and yellow, respectively.
Figure 2
Figure 2
Subcellular localization analysis of the PmARF6 protein. The control protein is represented by the 35S-GFP fusion. Merged 1: GFP + Auto; Merged 2: Merged 1 + Bright; GFP: green fluorescent protein; Auto: chlorophyll autofluorescence; scale bar 10 μm, represented by the red line.
Figure 3
Figure 3
qRT-PCR results of the PmARF6 gene. Male and Female represent male flowers and female flowers in P. massoniana, respectively. The error bars represent the standard deviations from three replicates.
Figure 4
Figure 4
Expression level and growth status of 35S::PmARF6 transgenic lines in Arabidopsis. (A) PmARF6 gene expression levels in WT Arabidopsis (control) and PmARF6-overexpressing Arabidopsis. WT: wild-type Arabidopsis; L1–L4: lines 1–4 for transgenic Arabidopsis. The error bars represent the standard deviations of three replicates. (B) The growth phenotypes of WT Arabidopsis (control) and PmARF6-overexpressing Arabidopsis.
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