The Copper Chaperone Protein Gene GmATX1 Promotes Seed Vigor and Seedling Tolerance under Heavy Metal and High Temperature and Humidity Stresses in Transgenic Arabidopsis

Abstract

Abiotic stresses such as high temperature, high humidity, and heavy metals are important factors that affect seed development and quality, and restrict yield in soybean. The ATX1-type copper chaperones are an important type of proteins that are used for maintaining intracellular copper ion homeostasis. In our previous study, a copper chaperone protein GmATX1 was identified in developing seeds of soybean under high temperature and humidity (HTH) stresses. In this study, the GmATX1 gene was isolated, and multiple alignment analysis showed that its encoding protein shared high sequence identities with other plant orthologues of copper chaperone proteins containing the HMA domain, and a conserved metal ion-binding site, CXXC. A subcellular localization assay indicated that GmATX1 was localized in the cell membrane and nucleus. An expression analysis indicated that GmATX1 was involved in seed development, and in response to HTH and heavy metal stresses in soybean. GmATX1-silent soybean seedlings were found to be more severely damaged than the control under HTH stress. Moreover, the silencing of GmATX1 reduced antioxidase activity and reactive oxygen species (ROS) scavenging ability in the seedling leaves. The overexpression of GmATX1 in Arabidopsis improved seed vigor and seedling tolerance, and enhanced antioxidase activity and ROS scavenging ability under HTH and heavy metal stresses. Our results indicated that GmATX1 could promote seed vigor and seedling tolerance to HTH and heavy metal stresses in transgenic Arabidopsis, and this promotion could be achieved by enhancing the antioxidase activity and ROS scavenging ability.

Phenylalanine Ammonia Lyase GmPAL1.1 Promotes Seed Vigor under High-Temperature and -Humidity Stress and Enhances Seed Germination under Salt and Drought Stress in Transgenic Arabidopsis

“…Seeds were transferred to H 2 DCFDA Buffer (1 mM H 2 DCFDA, 10 mM Hepes-NaOH, pH 5.7) and incubated in the dark for 30 min. It was observed by a laser confocal microscope (Zeiss LSM780, Jena, Germany) at 488 nm excitation and at 525 nm emission [ 36 ]. Each sample had at least three biological replicates.…”

“…The determination steps of antioxidant enzyme activity and MDA content of Arabidopsis thaliana leaves were conducted, as previously described [ 36 ]. In brief, the activity of superoxide dismutase (SOD) was determined by a nitrogen blue tetrazole method, while the activity of peroxidase (POD) was determined by a guaiacol method.…”