Abstract
Low temperature stress represents a significant abiotic stress factor affecting rice yields. While the structure and some of the functions of cell cycle protein-dependent protein kinase inhibitor (CKI) family proteins have been the subject of study, their relevance to cold tolerance in rice has been less investigated. In this study, we cloned OsEL2 (LOC_Os03g01740) and constructed anti-expression lines of this gene. The resulting lines exhibited significant cold sensitivity and displayed greater oxidative damage than wild type Nippobare (Nip). However, the activities of antioxidant enzymes, such as catalase (CAT), were significantly elevated in OsEL2-AX plants in comparison to Nip following exposure to 4 °C stress. RNA sequencing revealed the presence of 18,822 differential genes, with the majority of them being expressed with temporal specificity. The Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analysis revealed that a considerable number of differentially expressed genes (DEGs) are involved in the metabolism of amino acids, lipids, and terpenoids. Weighted gene co-expression network analysis (WGCNA) revealed a close relationship between the genes in the turquoise and light green modules and rice cold tolerance traits. These genes were predominantly enriched in terpene metabolism and the metabolism of various plant secondary metabolites, suggesting that OsEL2 influences rice cold tolerance through the metabolism of these two classes of substances. An analysis of the genes within these two modules using transcription factor (TF) enrichment and KEGG enrichment revealed that they are predominantly regulated by mitogen-activated protein kinase (MAPK) and ethylene signaling pathways. Furthermore, we found that tryptophan metabolism, phenylalanine metabolism, and monoterpene synthesis were enriched in down-regulated pathway enrichment analysis. In addition, we also found that the MAPK signaling pathway was enriched in the KEGG enrichment analysis of AX2 with Nip. The results demonstrate that anti-expression of OsEL2 is associated with a notable decline in rice tolerance to cold stress.