*3.6. Gene Ontology (GO) Analysis of Differentially Expressed Genes (DEGs)*

A GO enrichment analysis can be used to reveal the functional characteristics of differentially expressed genes (DEGs). GO terms are widely used to classify genes into the categories of cellular component (CC), molecular function (MF), and biological process (BP) [44]. As shown in Figure 5A, the GO enrichment analysis indicated that the DEGs were mostly enriched in biological processes (involving 31 BPs, *P*adjust ≤0.05) (Supplementary Table S4), and many of them were involved in the negative regulation of cell activities, such as the negative regulation of biological processes, cellular processes, and cell development, which indicated that DMM has negative toxic effects on Jurkat T cells. Moreover, many genes were also involved in the immune regulation of cell biological processes, such as lymphocyte activation and differentiation, T cell activation and differentiation, leukocyte differentiation, and negative regulation of the T cell apoptotic process. Most of the genes were downregulated in comparison to L4 vs. Ctr or M21 vs. Ctr (such as *RAG1*, *HDAC9*, *SOX4*, and *CD7*). However, only 12 genes (*AC138035.1*, *AP002990.1*, *BMP10*, *CHI3L2*, *DNTT*, *LINC01355*, *MME*, *MSH4*, *PAXIP1-AS1*, *PTPN3*, *SERPINB2*, and *TDRD9*) were upregulated, and four genes (*AL121594.1*, *CD40LG*, *EGR1*, and *SH3BP5*) were downregulated simultaneously in comparison to both L4 vs. Ctr and M21 vs. Ctr (Figure 5B). Among them, two important genes, *EGR1* and *CD40LG*, related to immune regulation were downregulated in both comparisons to L4 vs. Ctr and M21 vs. Ctr. Human CD40LG protein is a transmembrane protein and the ligand of CD40. It belongs to the tumor necrosis factor gene superfamily and is involved in immune-related pathways of breast cancer. The gene *CD40LG* plays critical roles in the regulation of the activation and differentiation of B cells

and the maturation of dendritic cells [45]. The gene *EGR1* (early growth response 1) is an important transcription factor that is widely expressed in many cell types and participates in important physiological processes of human cells [46]. A previous study indicated that *EGR1* serves as a tumor suppressor in cancers, such as prostate tumors and gastric tumors [47]. The downregulation of these genes may indicate that DMM exposure reduces the immune resistance of the body, even at a low concentration.

**Figure 4.** Venn diagram (**A**) and principal component analysis (PCA) for the differentially expressed genes (**B**). Hierarchical clustering analysis (HCA) of significantly differentially expressed genes (**C**) and statistics of the significantly differentially expressed genes (**D**) for each comparison.
