No abstract

Stress-associated proteins (SAPs), a group of zinc-finger-type proteins, have been identified as novel regulators of plant abiotic and biotic stresses. However, although they have been discovered in different plant species, their precise functional roles remain unclear. Here, we identified 14 SAP subfamily genes in the pepper genome. An investigation of the promoter regions of these genes for cis-regulatory elements associated with abiotic stress responses revealed the presence of multiple stress-related elements. Domain and phylogenetic analyses using the corresponding protein sequences revealed that the CaSAP genes can be classified into six groups (I–VI) and sorted into two broad types. Expression levels of the CaSAP genes were found to be differentially induced by low temperature, the dehydration stress, or exogenous abscisic acid. Group II and IV genes were highly induced by the low temperature and dehydration treatments, respectively. Moreover, subcellular localization analysis indicated that the proteins in these two groups are distributed in the nucleus, cytoplasm, and plasma membrane. Among the pepper plants silenced with the three identified group II CaSAP genes, the CA02g10410-silenced plants showed tolerance to low temperature, whereas the CA03g17080-silenced plants were found to have temperature-sensitive phenotypes. Interestingly, group IV CaSAP-silenced pepper plants showed drought-tolerant phenotypes. These findings contribute to a preliminary characterization of CaSAP genes and provide directions for future research on the biological role of CaSAPs in response to different abiotic stresses.

The phytohormone abscisic acid (ABA) plays a prominent role in various abiotic stress responses of plants.In the ABA-dependent osmotic stress response, SnRK2.6, one of the subclass III SnRK2 kinases, has been identified as playing a key role by phosphorylating and activating downstream genes. Although several modulatory proteins have been reported to be phosphorylated by SnRK2.6, the identities of the full spectrum of downstream targets have yet to be sufficiently established. In this study, we identified CaSAP14, a stress-associated protein in pepper (Capsicum annuum), as a downstream target of CaSnRK2.6. We elucidated the physical interaction between SnRK2.6 and CaSAP14, both in vitro and in vivo, and accordingly identified a C-terminal C2H2-type zinc finger domain of CaSAP14 as being important for their interaction. CaSAP14-silenced pepper plants showed dehydration-and high salt-sensitive phenotypes, whereas overexpression of CaSAP14 in Arabidopsis conferred tolerance to dehydration, high salinity, and mannitol treatment, with plants showing ABA-hypersensitive phenotypes. Furthermore, an in-gel kinase assay revealed that CaSnRK2.6 phosphorylates CaSAP14 in response to exogenous ABA, dehydration, and high-salinity stress. Collectively, these findings suggest that CaSAP14 is a direct substrate of CaSnRK2.6 and positively modulates dehydration-and high salinity-induced osmotic stress responses.

Abscisic acid (ABA) signalling triggers drought resistance mediated by SNF1‐related kinase 2s (SnRK2s), which transmits stress signals through the phosphorylation of several downstream factors. However, these kinases and their downstream targets remain elusive in pepper plants. This study aimed to isolate interacting partners of CaSnRK2.6, a homologue of Arabidopsis SnRK2.6/OST1. Among the candidate proteins, we identified a homeodomain‐leucine zipper (HD‐Zip) class II protein and named it CaHAT1 (Capsicum annuum homeobox ABA signalling related‐ transcription factor 1). CaHAT1‐silenced pepper and ‐overexpression (OE) transgenic Arabidopsis plants were generated to investigate the in vivo function of CaHAT1 in drought response. Following the application of drought stress, CaHAT1‐silenced pepper plants exhibited drought‐sensitive phenotypes with reduced ABA‐mediated stomatal closure and lower expression of stress‐responsive genes compared with control plants. In contrast, CaHAT1‐OE transgenic Arabidopsis plants showed the opposite phenotypes, including increased drought resistance and ABA sensitivity. CaHAT1, particularly its N‐terminal consensus sequences, was directly phosphorylated by CaSnRK2.6. Furthermore, CaSnRK2.6 kinase activity and CaSnRK2.6‐mediated CaHAT1 phosphorylation levels were enhanced by treatment with ABA and drought stress. Taken together, our results indicated that CaHAT1, which is the target protein of CaSnRK2.6, is a positive regulator of drought stress response. This study advances our understanding of CaHAT1–CaSnRK2.6 mediated defence mechanisms in pepper plants against drought stress.