Abstract
Environmental stress affects growth and development of crops, and reduces yield and quality of crops. To cope with environmental stressors, plants have sophisticated defense mechanisms, including the HSF/HSP pathway. Here, we identify the expression pattern of CaHSP16.4 in thermo-tolerant and thermo-sensitive pepper (Capsicum annuum L.) lines. Under heat stress, R9 thermo-tolerant line had higher CaHSP16.4 expression level than the B6 thermo-sensitive line. Under drought stress, expression pattern of CaHSP16.4 was dynamic. Initially, CaHSP16.4 was downregulated then CaHSP16.4 significantly increased. Subcellular localization assay showed that CaHSP16.4 localizes in cytoplasm and nucleus. In the R9 line, silencing of CaHSP16.4 resulted in a significant increase in malonaldehyde content and a significant reduction in total chlorophyll content, suggesting that silencing of CaHSP16.4 reduces heat and drought stresses tolerance. Overexpression of CaHSP16.4 enhances tolerance to heat stress in Arabidopsis. Under heat stress, the survival rate of CaHSP16.4 overexpression lines was significantly higher than wild type. Furthermore, under heat, drought, and combined stress conditions, the CaHSP16.4-overexpression lines had lower relative electrolytic leakage and malonaldehyde content, higher total chlorophyll content, and higher activity levels of superoxide dismutase, catalase, ascorbic acid peroxidase, and glutathione peroxidase compared to wild type. Furthermore, the expression levels of the stress response genes in the overexpression lines were higher than the wild type. These results indicate that the overexpression of CaHSP16.4 enhances the ability of reactive oxygen species scavenging under heat and drought stress.
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This work was supported through the funding from the National Natural Science Foundation of China (No. U1603102), National Key R&D Program of China (No. 2016YFD0101900), and the Independent Innovation Fund Project of Agricultural Science and Technology in Jiangsu (No.CX (17) 3040).
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LH and ZG designed the experiments. LH, GC, AK, AW, QY, and SY performed the research. LH drafted the manuscript. ZG revised the paper. ZG and DL contributed reagents/materials/analysis tools. All authors read and approved the final manuscript.
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Fig. S1
The silencing efficiency of CaHSP16.4. (a) Measurement of CaHSP16.4 expression levels in CaHSP16.4-TRV2 pepper lines by quantitative real-time PCR. The CaUBI3 gene was used as the internal control. Error bars represent standard deviation from three biological replicates. (b) Morphology of TRV2-CaHSP16.4, TRV2:00, and TRV2-CaPDS pepper lines under normal condition. (PNG 2855 kb)
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Fig. S2
Confirmation of CaHSP16.4-overexpressing Arabidopsis lines. (a) PCR confirmation of CaHSP16.4-overexpression Arabidopsis lines with genomic DNA as template. (b) Measurement of CaHSP16.4 expression levels in CaHSP16.4-overexpression Arabidopsis lines by quantitative real-time PCR. The AtActin2 gene was used as the internal control. Error bars represent standard deviation from three biological replicates. (PNG 727 kb)
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Table S1
Primer sequences used for quantitative real-time PCR analysis, subcellular localization, VIGS and overexpression. (DOCX 18 kb)
Table S2
Predicted subcellular localization of CaHSP16.4 based on online tools. (DOCX 16 kb)
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Huang, LJ., Cheng, GX., Khan, A. et al. CaHSP16.4, a small heat shock protein gene in pepper, is involved in heat and drought tolerance. Protoplasma 256, 39–51 (2019). https://doi.org/10.1007/s00709-018-1280-7
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DOI: https://doi.org/10.1007/s00709-018-1280-7