Limits...
A Non-specific Setaria italica Lipid Transfer Protein Gene Plays a Critical Role under Abiotic Stress

View Article: PubMed Central - PubMed

ABSTRACT

Lipid transfer proteins (LTPs) are a class of cysteine-rich soluble proteins having small molecular weights. LTPs participate in flower and seed development, cuticular wax deposition, also play important roles in pathogen and abiotic stress responses. A non-specific LTP gene (SiLTP) was isolated from a foxtail millet (Setaria italica) suppression subtractive hybridization library enriched for differentially expressed genes after abiotic stress treatments. A semi-quantitative reverse transcriptase PCR analysis showed that SiLTP was expressed in all foxtail millet tissues. Additionally, the SiLTP promoter drove GUS expression in root tips, stems, leaves, flowers, and siliques of transgenic Arabidopsis. Quantitative real-time PCR indicated that the SiLTP expression was induced by NaCl, polyethylene glycol, and abscisic acid (ABA). SiLTP was localized in the cytoplasm of tobacco leaf epidermal cells and maize protoplasts. The ectopic expression of SiLTP in tobacco resulted in higher levels of salt and drought tolerance than in the wild type (WT). To further assess the function of SiLTP, SiLTP overexpression (OE) and RNA interference (RNAi)-based transgenic foxtail millet were obtained. SiLTP-OE lines performed better under salt and drought stresses compared with WT plants. In contrast, the RNAi lines were much more sensitive to salt and drought compared than WT. Electrophoretic mobility shift assays and yeast one-hybrids indicated that the transcription factor ABA-responsive DRE-binding protein (SiARDP) could bind to the dehydration-responsive element of SiLTP promoter in vitro and in vivo, respectively. Moreover, the SiLTP expression levels were higher in SiARDP-OE plants compared than the WT. These results confirmed that SiLTP plays important roles in improving salt and drought stress tolerance of foxtail millet, and may partly be upregulated by SiARDP. SiLTP may provide an effective genetic resource for molecular breeding in crops to enhance salt and drought tolerance levels.

No MeSH data available.


Salt tolerance of SiLTP-OE and RNAi foxtail millet seedlings. The phenotypes of transgenic foxtail millet under a 100 mM NaCl treatment (A). Proline contents (B) and soluble sugar contents (C) were analyzed in WT and transgenic plants after watering for 14 and 21 days with NaCl, respectively. ∗, ∗∗, and ∗∗∗ indicate statistically significant differences at P < 0.05, P < 0.01, and P < 0.001 (Student’s t-test), respectively.
© Copyright Policy
Related In: Results  -  Collection

License
getmorefigures.php?uid=PMC5121218&req=5

Figure 9: Salt tolerance of SiLTP-OE and RNAi foxtail millet seedlings. The phenotypes of transgenic foxtail millet under a 100 mM NaCl treatment (A). Proline contents (B) and soluble sugar contents (C) were analyzed in WT and transgenic plants after watering for 14 and 21 days with NaCl, respectively. ∗, ∗∗, and ∗∗∗ indicate statistically significant differences at P < 0.05, P < 0.01, and P < 0.001 (Student’s t-test), respectively.

Mentions: Furthermore, 2-week-old seedlings of WT and transgenic plants were not irrigated for 10 days, and were then re-irrigated and grown under normal conditions for 3 days. The survival rates of the OE lines were higher than that of the WT, and the RNAi lines had much lower survival rates compared with the WT (Figures 8A,B). The proline and soluble sugar contents of the OE lines (especially the OE14 line) were higher than those of WT and RNAi lines after 10 days of drought treatment, and the RNAi lines had significantly lower soluble sugar contents than those of the WT and OE lines (Figures 8C,D). Moreover, the drought treatment was implemented during the reproductive stage of foxtail millet grown in a greenhouse. After 22 days of drought treatment, RNAi lines had visually retarded growth rates than WT and OE lines (Figure 8E). After 28 days of the drought treatment, the survival rate of RNAi lines significantly decreased compared with those of OE lines and the WT, and the OE line survival rates were much higher than those of WT and RNAi lines (Figures 8E,F). When 2-week-old seedlings of WT and transgenic plants were irrigated with water supplemented with 100 mM NaCl, none of the seedlings showed any significant differences after 14 days of treatment (Figure 9A). SiLTP-OE seedlings showed more tolerance compared with those of WT and RNAi lines after 21 days and 28 days of treatments (Figure 9A). After 21 days of treatment, the proline and soluble sugar contents of SiLTP-OE line 14 were significantly greater than those of the WT and RNAi lines (Figures 9B,C). The SiLTP-OE 69 line had a significantly greater proline content compared with the WT and RNAi lines, and its soluble sugar content was also greater, but not significantly (Figures 9B,C).


A Non-specific Setaria italica Lipid Transfer Protein Gene Plays a Critical Role under Abiotic Stress
Salt tolerance of SiLTP-OE and RNAi foxtail millet seedlings. The phenotypes of transgenic foxtail millet under a 100 mM NaCl treatment (A). Proline contents (B) and soluble sugar contents (C) were analyzed in WT and transgenic plants after watering for 14 and 21 days with NaCl, respectively. ∗, ∗∗, and ∗∗∗ indicate statistically significant differences at P < 0.05, P < 0.01, and P < 0.001 (Student’s t-test), respectively.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC5121218&req=5

Figure 9: Salt tolerance of SiLTP-OE and RNAi foxtail millet seedlings. The phenotypes of transgenic foxtail millet under a 100 mM NaCl treatment (A). Proline contents (B) and soluble sugar contents (C) were analyzed in WT and transgenic plants after watering for 14 and 21 days with NaCl, respectively. ∗, ∗∗, and ∗∗∗ indicate statistically significant differences at P < 0.05, P < 0.01, and P < 0.001 (Student’s t-test), respectively.
Mentions: Furthermore, 2-week-old seedlings of WT and transgenic plants were not irrigated for 10 days, and were then re-irrigated and grown under normal conditions for 3 days. The survival rates of the OE lines were higher than that of the WT, and the RNAi lines had much lower survival rates compared with the WT (Figures 8A,B). The proline and soluble sugar contents of the OE lines (especially the OE14 line) were higher than those of WT and RNAi lines after 10 days of drought treatment, and the RNAi lines had significantly lower soluble sugar contents than those of the WT and OE lines (Figures 8C,D). Moreover, the drought treatment was implemented during the reproductive stage of foxtail millet grown in a greenhouse. After 22 days of drought treatment, RNAi lines had visually retarded growth rates than WT and OE lines (Figure 8E). After 28 days of the drought treatment, the survival rate of RNAi lines significantly decreased compared with those of OE lines and the WT, and the OE line survival rates were much higher than those of WT and RNAi lines (Figures 8E,F). When 2-week-old seedlings of WT and transgenic plants were irrigated with water supplemented with 100 mM NaCl, none of the seedlings showed any significant differences after 14 days of treatment (Figure 9A). SiLTP-OE seedlings showed more tolerance compared with those of WT and RNAi lines after 21 days and 28 days of treatments (Figure 9A). After 21 days of treatment, the proline and soluble sugar contents of SiLTP-OE line 14 were significantly greater than those of the WT and RNAi lines (Figures 9B,C). The SiLTP-OE 69 line had a significantly greater proline content compared with the WT and RNAi lines, and its soluble sugar content was also greater, but not significantly (Figures 9B,C).

View Article: PubMed Central - PubMed

ABSTRACT

Lipid transfer proteins (LTPs) are a class of cysteine-rich soluble proteins having small molecular weights. LTPs participate in flower and seed development, cuticular wax deposition, also play important roles in pathogen and abiotic stress responses. A non-specific LTP gene (SiLTP) was isolated from a foxtail millet (Setaria italica) suppression subtractive hybridization library enriched for differentially expressed genes after abiotic stress treatments. A semi-quantitative reverse transcriptase PCR analysis showed that SiLTP was expressed in all foxtail millet tissues. Additionally, the SiLTP promoter drove GUS expression in root tips, stems, leaves, flowers, and siliques of transgenic Arabidopsis. Quantitative real-time PCR indicated that the SiLTP expression was induced by NaCl, polyethylene glycol, and abscisic acid (ABA). SiLTP was localized in the cytoplasm of tobacco leaf epidermal cells and maize protoplasts. The ectopic expression of SiLTP in tobacco resulted in higher levels of salt and drought tolerance than in the wild type (WT). To further assess the function of SiLTP, SiLTP overexpression (OE) and RNA interference (RNAi)-based transgenic foxtail millet were obtained. SiLTP-OE lines performed better under salt and drought stresses compared with WT plants. In contrast, the RNAi lines were much more sensitive to salt and drought compared than WT. Electrophoretic mobility shift assays and yeast one-hybrids indicated that the transcription factor ABA-responsive DRE-binding protein (SiARDP) could bind to the dehydration-responsive element of SiLTP promoter in vitro and in vivo, respectively. Moreover, the SiLTP expression levels were higher in SiARDP-OE plants compared than the WT. These results confirmed that SiLTP plays important roles in improving salt and drought stress tolerance of foxtail millet, and may partly be upregulated by SiARDP. SiLTP may provide an effective genetic resource for molecular breeding in crops to enhance salt and drought tolerance levels.

No MeSH data available.