m6A RNA methylation modulates IFN-γ-stimulated intestinal epithelial cell-intrinsic antiparasitic defense

N⁶-methyladenosine (m⁶A) RNA methylation is one of the most prevalent reversible post-transcriptional RNA modifications and has been recognized as a crucial regulator of host immune responses. Intestinal epithelial cells (IECs) constitute an important component of gastrointestinal mucosal immunity. Interferons (IFNs) play a central role in maintaining intestinal homeostasis, and m⁶A methylation status influences IFN-mediated cell-intrinsic defense. In this study, we investigated the potential role of m⁶A RNA modifications in IFN-γ-stimulated IEC-intrinsic defense. We observed significant alterations in the topology of the m⁶A mRNA methylome in murine IECs following IFN-γ stimulation. A subset of IFN-γ-stimulated immune gene transcripts exhibited increased m⁶A RNA methylation, including several members of the immunity-related GTPase family M (IRGM) genes. In addition, IFN-γ-responsive long non-coding RNAs may modulate the m⁶A methylation levels of multiple IFN-γ-stimulated immune transcripts. Enhanced m⁶A methylation of the Irgm2/3 transcripts was associated with strengthened cell-intrinsic defense against infection by the protozoan parasite Cryptosporidium. Notably, Cryptosporidium infection altered the host m⁶A mRNA methylome in IECs, thereby counteracting the IFN-γ-mediated defense response. Although the RNA levels of Irgm2/3 genes were upregulated, their m⁶A RNA methylation levels and protein expression were reduced in infected cells. This effect was associated with host delivery of dsRNAs derived from Cryptosporidium parvum virus 1, a virus harbored in the parasite. Collectively, our findings suggest that m⁶A methylation of RNA transcripts enhances IFN-γ-mediated IEC-intrinsic antiparasitic defense, while Cryptosporidium has evolved mechanisms to evade this response by suppressing m⁶A RNA methylation of IFN-γ-stimulated immune genes.