Recent advance in optical single-molecule detection of methylation modification and methyl-modifying enzymes

As the predominant epigenetic markers, methyl-modifying enzymes-catalyzed methylation modification widely participates in the regulation of cellular functions, and the dysregulation of methyl-modifying enzyme activity is closely related with various malignant tumors, autoimmune disorders, and diabetes. Consequently, methylation modification and methyl-modifying enzymes have become valuable biomarkers for clinical disease diagnosis. Single-molecule fluorescent detection has distinct advantages of minimum sample consumption, excellent sensitivity, and visualization capability. However, there is no review about single-molecule detection of methylation modification and methyl-modifying enzymes so far. Herein, we summarize the single-molecule detection-based strategies for the measurement of methylation modification and methyl-modifying enzymes in the last five years (2019–2023), and focus on five categories of methylation modification and methyl-modifying enzymes including DNA/RNA methylation modifications, DNA methyltransferases, DNA demethylases, RNA demethylases, and RNA methyltransferases. We summarize the working principles and applications of these strategies, and discuss the challenges and prospects in this field.