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.

SBP-box (Squamosa-promoter binding protein) genes are a type of plant-specific transcription factor and play important roles in plant growth, signal transduction and stress response. However, little is known about the SBP-box genes in pepper (CaSBP), especially in the process of Phytophthora capsici infection. In this study, a novel gene (CaSBP12) was selected from the CaSBP gene family, which was isolated from the pepper genome database in our previous study. The CaSBP12 gene was located in the nucleus of the cell and its silencing in the pepper plant enhanced the defense response against Phytophthora capsici infection. After inoculation with Phytophthora capsici, the root activity of the CaSBP12-silenced plants is compared to control plants, while malondialdehyde (MDA) content is compared viceversa. Additionally, the expression of defense related genes (CaPO1, CaSAR8.2, CaBPR1, and CaDEF1) in the silenced plants were induced to different degrees and the peak of CaSAR8.2 and CaBPR1 were higher than that of CaDEF1. The CaSBP12 over-expressed Nicotiana benthamiana plants were more susceptible to Phytophthora capsici infection with higher EC (electrical conductivity) and MDA contents as compared to the wild-type. The relative expression of defense related genes (NbDEF, NbNPR1, NbPR1a, and NbPR1b) in transgenic and wild-type Nicotiana benthamiana plants were induced, especially the NbPR1a and NbPR1b. In conclusion, these results indicate that CaSBP12 gene negative regulates the defense response against Phytophthora capsici infection which suggests their potentially significant role in plant defense. To our knowledge, this is the first report on CaSBP gene which negative regulate defense response.

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