Lipoxygenases (LOXs) play significant roles in abiotic stress responses, and identification of LOX gene promoter function can make an important contribution to elucidating resistance mechanisms. Here, we cloned the CmLOX08 promoter of melon (Cucumis melo) and identified the main promoter regions regulating transcription in response to signalling molecules and abiotic stresses. The 2054-bp promoter region of CmLOX08 from melon leaves was cloned, and bioinformatic analysis revealed that it harbours numerous cis-regulatory elements associated with signalling molecules and abiotic stress. Five 5′-deletion fragments obtained from the CmLOX08 promoter—2054 (LP1), 1639 (LP2), 1284 (LP3), 1047 (LP4), and 418 bp (LP5)—were fused with a GUS reporter gene and used for tobacco transient assays. Deletion analysis revealed that in response to abscisic acid, salicylic acid, and hydrogen peroxide, the GUS activity of LP1 was significantly higher than that of the mock-treated control and LP2, indicating that the − 2054- to − 1639-bp region positively regulates expression induced by these signalling molecules. However, no deletion fragment GUS activity was induced by methyl jasmonate. In response to salt, drought, and wounding treatments, LP1, LP2, and LP4 promoted significantly higher GUS expression compared with the control. Among all deletion fragments, LP4 showed the highest GUS expression, indicating that − 1047 to − 1 bp is the major region regulating promoter activity and that the − 1047 to − 418-bp region positively regulates expression induced by salt, drought, and wounding, whereas the − 1284 to − 1047-bp region is a negative regulatory segment. Interestingly, although the GUS activity of LP1 and LP2 was not affected by temperature changes, that of LP3 was significantly induced by heat, indicating that the − 1284- to − 1-bp region is a core sequence responding to heat and the − 2054- to − 1284-bp region negatively regulates expression induced by heat. Similarly, the − 1047- to − 1-bp region is the main sequence responding to cold, whereas the − 2054- to − 1047-bp region negatively regulates expression induced by cold. We cloned the CmLOX08 promoter and demonstrated that it is a signalling molecule/stress-inducible promoter. Furthermore, we identified core and positive/negative regulatory regions responding to three signalling molecules and five abiotic stresses.

NAC transcription factors (TFs) play important roles in plants’ responses to abiotic stresses and developmental processes, including leaf senescence. Oriental melon (Cucumis melo var. makuwa Makino) is an important vegetable crop in China and eastern Asia countries. However, little is known about the functions of the melon NAC family members. In this study, a phylogenetic tree was constructed to show that CmNAC60 and the senescence regulator AtNAP were in the same cluster, which implied that CmNAC60 might be a NAC related to leaf senescence. The expression analysis of CmNAC60 in different melon organs showed that the expression of CmNAC60 was highest in the male flowers and lowest in the hypocotyl. In addition, the expression level of CmNAC60 in the senescing leaves was significantly higher than in the non-senescing leaves. Similarly, the expression level of CmNAC60 in the dark-treated leaves was significantly higher than in the untreated leaves. Furthermore, the subcellular localization and transcriptional activation assays indicated that CmNAC60 was a nucleus localized NAC transcription factor with a C-terminal transactivation domain. An analysis of the tissue specific expression showed that the promoter of CmNAC60 may contain cis-acting regulatory elements responsive to leaf senescence. CmNAC60 overexpressing lines of Arabidopsis showed a precocious senescence compared with the wild type (WT). Collectively, our results showed that CmNAC60 was associated with leaf senescence, and could be potentially utilized in molecular breeding to improve melon yield or to extend the postharvest shelf life by delaying leaf senescence.

Thioredoxin z (TRX z) plays a significant role in chloroplast development by regulating the transcription of chloroplast genes. In this study, we identified a pentatricopeptide repeat (PPR) protein, rice albino seedling-lethal (RAS), that interacts with OsTRX z. This interaction was initially discovered by using a yeast two-hybrid (Y2H) screening technique and was further validated through Y2H and bimolecular fluorescence complementation (BiFC) experiments. RAS contains 16 PPR motifs and features a small MutS-related (SMR) domain at its C-terminus. CRISPR/Cas9-generated ras mutants exhibited an albino seedling-lethal phenotype characterized by abnormal chloroplast structures and a significantly reduced chlorophyll content. RAS localizes to the chloroplast and is predominantly expressed in young leaves. Mutations in RAS affect RNA editing at the rpl2, rps14, and ndhA sites, as well as RNA splicing at the rpl2, atpF, and ndhA transcripts within the chloroplast. Furthermore, the expression levels of genes associated with chloroplast formation are altered in the ras mutant. Both OsTRX z and RAS were found to interact with chloroplast signal recognition particle (cpSRP) proteins, indicating that their proper localization within the chloroplast may be dependent on the SRP pathway. Collectively, our findings highlight the critical role of RAS in chloroplast development, as it is involved in RNA processing and the regulation of chloroplast gene expression.