Effective RNAi-Mediated Silencing of the Mismatch Repair MSH2 Gene Induces Sterility of Tomato Plants but Not an Increase in Meiotic Recombination.

Svetlana R Strelnikova,Roman A Komakhin,Anastasiya A Krinitsina

doi:10.3390/genes12081167

Abstract

In plant breeding, the ability to manipulate meiotic recombination aids in the efficient construction of new allelic compositions of chromosomes and facilitates gene transfer from wild relatives of crop plants. The DNA mismatch repair system antagonizes meiotic recombination. In this research, a trial was conducted to evaluate transgenic tomato plants carrying an RNA interference (RNAi) construct designed to inhibit the expression of the mismatch repair MSH2 gene. To drive the RNAi construct, we used either a pro-SmAMP2 promoter from Stellaria media ANTIMICROBIAL PEPTIDE2 or a Cauliflower mosaic virus 35S promoter (CaMV35S). The results of real-time PCR showed that, with a 16 h light/8 h dark photoperiod, MSH2-RNAi tomato transgenic plants exhibited MSH2 gene transcript contents ranging from 0% to 3% in the leaves, relative to untransformed controls. However, with this lighting mode, the MSH2-RNAi transgenic plants grew slowly, flowered poorly, and did not form seed sets. During cultivation with a 12 h light/12 h dark photoperiod, MSH2-RNAi transgenic plants exhibited MSH2 gene transcript contents ranging from 3% to 42%, relative to untransformed controls. Under these conditions, F1 hybrid seed sets formed for most of the MSH2-RNAi transgenic plants with the RNAi construct driven by the CaMV35S promoter, and for one transformant with the RNAi construct driven by the pro-SmAMP2 promoter. Under conditions of a 12 h light/12 h dark photoperiod, most of the F1 transgenic hybrids showed MSH2 gene transcript contents ranging from 3% to 34% and formed F2 offspring sets, which made it possible to assess the meiotic recombination frequency. We showed that the effective inhibition of MSH2 in MSH2-RNAi tomato transgenic plants is not associated with an increase in meiotic recombination compared to the control, but it stimulates the sterility of plants. It was established that the expression of the MSH2 gene in tomato plants is about 50 times higher with a 12 h light/12 h dark than with a 16 h light/8 h dark photoperiod. It is discussed that, in Solanum lycopersicum tomato plants, which are not sensitive to the day length for flowering, changing the lighting time may be a means of controlling the meiotic recombination frequency within certain limits.

Highlights

We included in our research eight and nine independent MSH2-RNA interference (RNAi) tomato transgenic plants from T0 pSmAMP2-antM2 and T0 p35S-antM2 groups, respectively, with at least 97% RNAi silencing of the MSH2 transcript
Abnormalities in plant growth, fruit formation, and seed set were found in all MSH2-RNAi plants under “long-day” conditions
It was previously noted that the knockout mutation of the MSH2 gene in Arabidopsis plants leads to the intensive accumulation of mutations in the genome over a number of generations, partial fertility loss, and a small seed set [16,33]

Summary

Introduction

Meiosis is a fundamental process shared by most sexually reproducing eukaryotic organisms. It generates genetic diversity through three principal mechanisms: (i) pairs of homologous chromosomes are independently sorted into haploid gametes; (ii) pairs of homologous chromosomes reciprocally exchange (crossing over) chromosome segments, altering the association of maternal and paternal alleles; and (iii) gene conversion occurs during homologous recombination [1]. In plant breeding, gaining control over the processes which alter crossing-over positions on chromosomes and increase the crossover frequency allow the more efficient construction of new allelic compositions of chromosomes for crop plants [2,3,4]. The ability to manipulate meiotic crossing over may be useful to facilitate the transmission of allelic genes from wild relatives to cultivated plants [5]

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