Exogenous Promoter Triggers APETALA3 Silencing through RNA-Directed DNA Methylation Pathway in Arabidopsis.

Benqi Wang,Lei Chu,Xue Jing,Jian Guo,Bin Yi,Huadong Wang,Jie Liu

doi:10.3390/ijms20184478

Benqi Wang, Lei Chu + Show 5 more

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https://doi.org/10.3390/ijms20184478

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Abstract

The development of floral organs plays a vital role in plant reproduction. In our research, the APETALA3 (AP3) promoter-transgenic lines showed abnormal developmental phenotypes in stamens and petals. The aim of this study is to understand the molecular mechanisms of the morphological defects in transgenic plants. By performing transgenic analysis, it was found that the AP3-promoted genes and the vector had no relation to the morphological defects. Then, we performed the expression analysis of the class A, B, and C genes. A dramatic reduction of transcript levels of class B genes (AP3 and PISTILLATA) was observed. Additionally, we also analyzed the methylation of the promoters of class B genes and found that the promoter of AP3 was hypermethylated. Furthermore, combining mutations in rdr2-2, drm1/2, and nrpd1b-11 with the AP3-silencing lines rescued the abnormal development of stamens and petals. The expression of AP3 was reactivated and the methylation level of AP3 promoter was also reduced in RdDM-defective AP3-silencing lines. Our results showed that the RdDM pathway contributed to the transcriptional silencing in the transgenic AP3-silencing lines. Moreover, the results revealed that fact that the exogenous fragment of a promoter could trigger the methylation of homologous endogenous sequences, which may be ubiquitous in transgenic plants.

Highlights

T-DNA insertion mutants of Arabidopsis are invaluable resource for studies of gene functions. [1]
The results show that the expression of AP3 is correlated with the hypermethylation of the AP3 promoter mediated by the RdDM pathway in the transgenic plants of Arabidopsis
The results suggest that DNA methylation of PI promoter was not observed at all three cytosine contexts in M17 and Col-0 (Figure 4B), which is opposite to the level of PI expression

Summary

Introduction

T-DNA insertion mutants of Arabidopsis are invaluable resource for studies of gene functions. [1]. T-DNA insertion mutants of Arabidopsis are invaluable resource for studies of gene functions. The silencing of 35S promoter-driven transgenes might occur in T-DNA (SALK, GABI, and FLAG) insertion mutants [2,3]. Studies have shown that some T-DNA transgene silencing of the 35S promoter insertion is short-interfering RNAs (siRNA)-mediated [2]. The expression of a target gene or an endogenous gene is inhibited by the transfer of a foreign gene into a plant, but usually only affects the transferred gene and its endogenous gene and does not affect the expression of other genes. Plant transgenic silencing was first discovered in 1990 [4]. As the research has progressed, it is found that transgene silencing is universal and can even affect plant development

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