Speckled Protein 140 (SP140) belongs to the Speckled Protein family of chromatin readers, acting as a central interpreter of the epigenome that facilitates normal immunological responses. SP140 contains chromatin-related modules, a PHD-BRD domain that recognizes histone modifications, a CARD domain mediating protein multimerization, and a SAND domain previously purposed to mediate DNA binding. These domains suggest that SP140 can bind to DNA directly; however, the mechanism that underlies the chromatin association of SP140 remains undefined. Here, I demonstrate that SP140 SAND preferentially binds nonmethylated CpG dinucleotides, and its binding is abrogated by DNA methylation. This binding characteristic leads to the association of SP140 with nonmethylated CpG islands within gene promoters, underlying the recruitment mechanism of SP140 for regulating immune-related genes. A conserved DNA-binding KNWR motif is also identified in SP140 SAND, and mutation of the critical amino acids within its KNWR motif completely abrogates its DNA-binding ability. Taken together, this work revealed a previously unrecognized chromatin recruitment mechanism of SP140 by using a combinatorial approach, offering a new perspective on the role of SP140 in immune regulation.