Legume Genomics and Genetics 2025, Vol.16, No.5, 234-244 http://cropscipublisher.com/index.php/lgg 237 3.2 Potential regulatory network of the ICE-CBF-COR pathway in adzuki bean The well-known cold regulation module in Arabidopsis thaliana - the ICE-CBF-COR pathway - may not be "groundless" in adzuki beans either. At the ICE beginning, once a cold signal is introduced, it is responsible for activating the expression of CBF. The CBF will continue to drive the work of the COR gene. This series of "relays" will enable cells to enhance antioxidant capacity, stabilize membrane structure, and even reduce the damage caused by dehydration in a low-temperature environment (Ding et al., 2022). However, the performance of the ICE protein itself is not static. It is controlled by multiple kinases and ubiquitin systems. That is to say, the time and intensity of its "activation" also vary in response to the duration of external cold stimulation. Although our understanding of this system in adzuki beans is not yet deep enough, leguminous plants generally possess similar regulatory potential. Current studies have provided some evidence for its existence (Kidokoro et al., 2022). 3.3 Other cold-responsive transcription factors (e.g., MYB, bZIP, NAC) and their functions CBF is indeed a "leading player", but it is not the only "trader". Transcription factor families like MYB, bZIP and NAC are also involved in different stages of cold adaptation to varying degrees. MYB, especially those members that can collaborate with bHLH proteins, often participate in the regulation of antioxidant substances such as anthocyanins (Mehrotra et al., 2020). The bZIP family, on the other hand, is more involved in the restart of transcription programs or the establishment of stress memories. NAC transcription factors have received the most attention in recent years. They can not only independently regulate cold-induced genes but also may "cross forces" with the CBF pathway, making them increasingly important in regulating cold responses (Diao et al., 2020; Abdullah et al., 2022). Overall, these "supporting roles" may not play as concentrated a role as CBF, but within the entire network, their regulatory "methods" are more diverse. 4 Epigenetic and Non-Coding RNA Regulatory Mechanisms 4.1 Regulation of cold-responsive genes by DNA methylation and histone modifications Not all genes will react immediately in a cold environment. Sometimes, plants need a more "cautious" regulatory approach to determine when genes are turned on and off. This "switch" mechanism involves DNA methylation and histone modification. In the study of Arabidopsis thaliana, long non-coding Rnas (lncrnas) such as SVALKA recruit multi-comb complex PRC2 to demethylate histones of cold-response genes like CBF3, resulting in tighter chromatin and greater difficulty in transcription (Kiger and Schroeder, 2024). However, this regulation is not static. Under prolonged low temperatures, it is actually dynamically adjusted - neither keeping the cold response on all the time nor turning it off too early. This mechanism is very likely to have a similar "universal version" in leguminous plants, such as adzuki beans. 4.2 Post-transcriptional regulation mediated by miRNAs under cold stress Not all cold stress responses occur at the "starting point" of gene expression. The "behind-the-scenes role" of miRNA emerges in the post-transcriptional stage. It does not activate genes but selectively "silences" some mRNA that has already been produced. Cold response mirnas such as miR397, miR408, miR394, which have long been discovered in Arabidopsis thaliana, as well as miR156, miR319 and miR528 in rice, have different functions - some affect hormone signaling, some regulate antioxidation, and some intervene in metabolism (Huo et al., 2021). High-throughput sequencing has gradually clarified these regulatory networks. Not only in model plants, but also in legumes and grains, a large number of cold-related mirnas have been discovered. Although adzuki beans have received less research, based on the existing data, they may also be equipped with a similar miRNA regulatory system. 4.3 Potential roles of lncRNAs and circRNAs in cold adaptation Long non-coding Rnas (lncrnas) and circular Rnas (circrnas) may not sound like the main characters, but at low temperatures, they could be the "tuners" maintaining system stability. Take SVALKA as an example. It is no mere decoration in Arabidopsis thaliana - it regulates the activity of CBF-like transcription factors by interfering with transcription, recruiting chromatin modification complexes, and even influencing miRNA pathways. Furthermore, lncRNA can also act as ceRNA to competitively "adsorb" miRNA, indirectly allowing cold response genes to
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