Overexpression of GmNFYA5 confers drought tolerance to transgenic Arabidopsis and soybean plants

Abstract

Background: Crop productivity is challenged by abiotic stresses, among which drought stress is the most common. NF-Y genes, especially NF-YA genes, regulate tolerance to abiotic stress. Results: Soybean NF-Y gene GmNFYA5 was identified to have the highest transcript level among all 21 NF-YA genes in soybean (Glycine max L.) under drought stress. Drought-induced transcript of GmNFYA5 was suppressed by the ABA synthesis inhibitor naproxen (NAP). GmNFYA5 transcript was detected in various tissues at vegetative and reproductive growth stages with higher levels in roots and leaves than in other tissues, which was consist with the GmNFYA5 promoter: GUS fusion assay. Overexpression of GmNFYA5 in transgenic Arabidopsis plants caused enhanced drought tolerance in seedlings by decreasing stomatal aperture and water loss from leaves. Overexpression and suppression of GmNFYA5 in soybean resulted in increased and decreased drought tolerance, respectively, relative to plants with an empty vector (EV). Transcript levels of ABA-dependent genes (ABI2, ABI3, NCED3, LEA3, RD29A, P5CS1, GmWRKY46, GmNCED2 and GmbZIP1) and ABA-independent genes (DREB1A, DREB2A, DREB2B, GmDREB1, GmDREB2 and GmDREB3) in transgenic plants overexpressing GmNFYA5 were higher than those of wild-type plants under drought stress; suppression of GmNFYA5 transcript produced opposite results. GmNFYA5 probably regulated the transcript abundance of GmDREB2 and GmbZIP1 by binding to the promoters in vivo. Conclusions: Our results suggested that overexpression of GmNFYA5 improved drought tolerance in soybean via both ABA-dependent and ABA-independent pathways.

GmNFYA13 Improves Salt and Drought Tolerance in Transgenic Soybean Plants

“…Merr.) cultivar Williams 82 were grown in plastic pots (14 cm diameter, 15 cm depth) containing nutrient soil and grown for 20 days in a tissue culture room at 28/18 • C day/night with a 14-h photoperiod and 70% relative humidity as described previously (Ma et al, 2020), which are considered as normal conditions for soybean plants. The seedlings were used to assess gene expression patterns under several treatments.…”

“…For H 2 O and salt treatment, the seedlings were subjected to H 2 O and 200 mM NaCl solution for 12 h; for drought treatment, the seedlings were pulled out from soil and placed on a plastic plate for 12 h; for ABA and H 2 O 2 treatment, the roots were immersed in 100 µM ABA and 10 mM H 2 O 2 for 12 h. The leaves under various treatments were harvested at 0, 1, 2, 4, 8, and 12 h and frozen at −80 • C for isolation of RNA. To test whether an ABA synthesis inhibitor (tungstate) could affect salt-induced GmNFYA13 transcripts, two detached leaves of soybean plants were placed into ddH 2 O to eliminate the effect of wound stress for 1 h, and then transferred into ddH 2 O and 2 mM tungstate for 3 h, followed by being subjected to 200 mM NaCl solution for 1 h. To determine whether tungstate could suppress the drought-induced expression level of GmNFYA13, the "drought + tungstate" treatment was performed as described previously (Ma et al, 2020) and 1 mM naproxen was replaced with 2 mM tungstate. The leaves immersed in ddH 2 O were used as the control.…”

“…To evaluate the transcription levels of GmNFYA13 in different tissues, the tissues of 20-day-old and 50-day-old soybean plants were collected and stored as described previously (Ma et al, 2020). All of the experiments were performed in a tissue culture room at 28/18 • C day/night with a 14-h photoperiod and 70% relative humidity.…”

“…Roots of 35S:GmNFYA13 plants could express GFP-GmNFYA13 fusion protein. Roots of 35S:GmNFYA13 and WT Arabidopsis plants were detached and observed using a confocal laser scanning microscope (Zeiss, LSIM700) as described previously (Liu et al, 2013;Ma et al, 2020).…”

“…The 2000-bp DNA fragment upstream of cDNA was amplified to obtain the GmNFYA13 promoter using PrimerSTAR Max DNA polymerase (Takara, Japan) with the primer set 5 -GGACACGTGTCTCGATCTTCG-3 and 5 -CTTAGCACAACTTTTCCCTAT-3 . The products were cloned into a modified pCAMBIA1305 vector to generate the pCAMBIA1305:P GmNFYA13 fusion construct with the forward primer 5 -CCATGATTACGAATTCGGACACGTGTCTCGATCTTCG-3 and reverse primer 5 -CTCAGATCTACCATGGCTTAGCACAA CTTTTCCCTAT-3 as previously described (Ma et al, 2020). The A. tumefaciens strain GV3101 harboring pCAMBIA1305:P GmNFYA13 was used to infect Arabidopsis flowers.…”