Triticeae Genomics and Genetics, 2025, Vol.16, No.6, 262-268 http://cropscipublisher.com/index.php/tgg 265 4.2 GO annotation and KEGG pathway enrichment analysis After identifying the relevant modules, it is not immediately known what they are doing. At this point, one has to rely on annotation tools like GO and KEGG. The results were not too unexpected either: Many module genes were concentrated in the categories of hormone signal transduction, developmental regulation, and transcription factor activity, which were closely related to panicle development (Li et al., 2019; 2022). These enriched information are equivalent to "labeling" the module. Although they do not represent all the functions, they can at least provide a starting point for interpretation. 4.3 Identification of key hub genes within modules and construction of regulatory networks The genes in the module are not equal. Some genes have particularly large "social circles" and are related to many other genes, and are thus called hub genes. Screening out these hubs is to then draw the network diagram to see if they are the core nodes on the regulatory chain (Wang et al., 2017; VanGessel et al., 2022). This kind of network analysis not only helps us understand how genes check and balance each other or cooperate, but also makes it convenient to pick out candidate targets worth using for transgenic or editing experiments. For those engaged in variety improvement, these genes might be the most valuable breakthrough points. 5 Analysis of Transcription Factors and Regulatory Networks 5.1 Identification of transcription factor families associated with spike development modules (e.g., MADS-box, bHLH) In fact, in the panicle development module, what was first noticed was not a specific gene, but some clearly concentrated families of transcription factors - familiar faces like MADS-box and bHLH, which account for a large proportion in many modules. They are not the first time to be associated with the development of floral organs and meristem, but this co-expression network analysis has still made their roles somewhat clearer. Especially the MADS-box genes, during the gradual development of the spike, their activation sequence is very regular (VanGessel et al., 2022; Lin et al., 2024). Although such families have long been studied, their continuous screening out in the spike-related modules indicates that their main regulatory role still cannot be ignored. 5.2 Expression pattern analysis between TFs and their target genes within modules It's not enough to just look at the transcription factors; you also need to see who they partner with. By analyzing the expression patterns of transcription factors and their potential target genes, researchers found that many of them were simultaneously upregulated at the same developmental time point. This rhythmic coordination suggests that they may have a "partner" relationship in spike initiation and differentiation (Xu et al., 2025). Interestingly, these pairings are not scattered but concentrated in specific modules, indicating that the regulatory programs within these modules are very tight. Compared with the spatio-temporal expression data, this phenomenon of expression synchronization is also more convincing, helping us gradually piece together the regulatory map of panicle development. 5.3 Signal transduction pathways and regulatory mechanisms mediated by TFs But the story of transcription factors is far more than just "up-regulation and down-regulation". In many cases, they are also caught between hormone signals and gene expression, playing the role of "translators". Familiar plant hormones such as auxin and cytokinin often exert their regulatory effects through TF (Li et al., 2019). Of course, the pathways mediated by different TFS are not exactly the same. Some control the morphological construction of the spike, while others may be more inclined towards organ-level transformation. All this information, when put together, helps explain how the final structure of the spike was gradually "determined" and also points out possible genetic targets for subsequent yield improvements. 6 Case Study: Empirical Validation of Key Modules and Genes 6.1 Correlation analysis of the “turquoise” module with spike length regulation In the research, panicle length is regarded as one of the main factors affecting wheat yield. However, not all expression modules are related to this trait. The peculiarity of the "turquoise" module lies in that multiple genes within it exhibit expression patterns significantly related to panicle length in different wheat varieties (Wei et al.,
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