A small RNA of miR2119b from soybean CMS line acts as a negative regulator of male fertility in transgenic Arabidopsis

Abstract

The miR2119 is involved in the growth, development and abiotic stress response of some legumes, including Medicago truncatula, Phaseolus vulgaris and soybean (Glycine max (L.) Merr.). Our previous small RNA sequencing analysis showed that miR2119b was up-regulated in the flower buds of soybean cytoplasmic male sterile (CMS) line compared with its maintainer line, but the role and mechanism of miR2119b in the regulation of soybean male fertility are still unclear. In this study, the gma-miR2119b and its target gene alcohol dehydrogenase 1.3b (ADH1.3b) were characterized and found to be highly expressed in the flowers of soybean CMS line and its maintainer. Transgenic Arabidopsis plants overexpressing gma-miR2119b exhibit male fertility abnormalities, including pollen fertility and germination rate decreased. Enzyme activity detection found the ADH and catalase (CAT) enzyme activities in inflorescence of gma-miR2119b overexpressed plants were lower than those of wild-type. Bioinformatics and gene expression analysis showed that gma-miR2119b/GmADH1.3b module was responsive to high temperature (HT) stress during flowering. After HT stress, the gma-miR2119b overexpressed plants showed male sterility, including shorter filament, sterile pollen, indehiscent anther and non seed. Moreover, some key genes involved in HT response and reactive oxygen species (ROS) signal regulation pathway, including heat shock protein70, galactinol synthase 1 and CAT, showed down-regulated expression in transgenic plants under HT stress, suggesting that gma-miR2119b regulates male fertility via HT-ROS signaling pathway under HT stress. It was speculated that the gma-miR2119b acted as a negative regulator of male fertility in plants by regulating ADH1, HT-induced and ROS scavenging genes expression.