CpCML subverts host PI3K/AKT-NF-κB signaling and autophagic flux via direct interaction with SUGT1 to facilitate Cryptosporidium parvum infection.

Cryptosporidium parvum is a water and foodborne apicomplexan pathogen that vigorously disrupts host signaling networks to enable intracellular colonization. However, the parasite-encoded effectors responsible for this modulation and the specific host signaling cascades they involve is still unexplored. In this study, we identify CpCML, a calmodulin-like protein encoded by C. parvum, as a perilous virulence effector that modulates host PI3K-NF-κB signaling and autophagic flux through direct interaction with the host protein SUGT1. Biochemical and cellular analyses reveal that CpCML binds SUGT1 with high specificity, inducing its redistribution from the cytoplasm to the nucleus, a shift that fundamentally reprograms host signaling, and thereby promoting nuclear enrichment. Expression of CpCML enhances phosphorylation of AKT and subsequent activation of NF-κB without changing total NF-κB or AKT protein levels, indicative of post-translational modulation of signaling activity rather than changes in protein expression. Inhibition of PI3K abrogates CpCML-induced NF-κB activation and autophagy-related responses, establishing a functional dependency of these outcomes on PI3K signaling. Mechanistic dissection further reveals that SUGT1 serves as a pivotal node: siRNA-mediated depletion of SUGT1 assists NF-κB activation and promotes the accumulation of autophagy-associated markers (LC3-II and p62), indicative of impaired autophagic flux, which in turn helps intracellular parasite survival; on the contrary, SUGT1 overexpression restores autophagic flux and significantly diminishes parasite burden. Together, these findings outline a CpCML-SUGT1-PI3K-NF-κB signaling axis that integrates host signaling and autophagy responses during C. parvum infection, presents a mechanistic framework for understanding how this obligate intracellular parasite preserved conserved cellular pathways to confirm its survival and replication.