Defining the role of ANKEF1 in ciliary function and host defense against influenza virus infection

The National Institute of Allergy and Infectious Diseases (NIAID) awarded $856,791 under the Allergy and Infectious Diseases Research program (CFDA 93.855) to the Icahn School of Medicine at Mount Sinai on September 12, 2025, for a Project Grant focused on defining the role of ANKEF1 (ankyrin repeat and EF-hand domain containing gene 1) in ciliary function and host defense against influenza virus infection. The award supports investigative research addressing why some otherwise healthy individuals, particularly children, develop severe influenza complications such as pneumonia, acute respiratory distress syndrome, or encephalitis. The research will utilize novel mouse models, human induced pluripotent stem cell (iPSC)-derived airway epithelial cultures, and advanced multi-omics technologies to examine how ANKEF1 deficiency impairs mucociliary clearance—the respiratory tract's primary defense mechanism against pathogens—and how persistent viral presence may trigger exaggerated inflammatory responses that increase susceptibility to influenza-associated encephalitis.

The project incorporates collaborative sub-awards to Rockefeller University and Boston University Medical Campus that expand the research scope through specialized capabilities. Rockefeller University will conduct genetic analysis of the patient's family through whole exome/whole genome sequencing to confirm genotype-phenotype associations and generate iPSCs from fibroblast samples for cellular mechanism investigation. Boston University Medical Campus will perform CRISPR/Cas9 gene-editing of iPSC-derived basal cells to introduce mutations of interest and differentiate the edited cells into mucociliary airway epithelium for phenotypic analysis. The project period extends through August 31, 2026, with work performed in New York, NY.

Abstract

Most influenza cases resolve without complications, but some progress to serious conditions like pneumonia, acute respiratory distress syndrome, or encephalitis. Genetic predispositions, such as inborn errors of immunity, are known factors that contribute to these severe outcomes, yet the underlying mechanisms in otherwise healthy individuals, particularly children, remain unclear. This proposal focuses on ANKEF1, a ciliary protein identified as having a loss-of-function mutation in a child who succumbed to influenza-associated rhombencephalitis-a rare and severe complication characterized by inflammation of the brainstem that can occur in some severe cases of influenza-associated encephalitis (IAE). Our preliminary findings indicate that ANKEF1 is critical for ciliary motility in airway epithelial cells, which is essential for effective mucociliary clearance (MCC). In mouse models, ANKEF1 deficiency leads to impaired MCC, reducing the ability to clear pathogens from the respiratory tract and leading to increased susceptibility to influenza infection, higher morbidity, and greater mortality. We hypothesize that persistent viral presence in the airways due to defective MCC triggers an exaggerated inflammatory response, which elevates the risk of IAE through increased cytokine production and disruption of brain barrier integrity in this proposal, we will leverage novel mouse models, human ipsc-derived airway epithelial cultures, and advanced multi-omics technologies to define how ANKEF1 regulates ciliary function and host defense. Aim 1 will investigate how ANKEF1 deficiency alters ciliary beat frequency, coordination, and motility in the airways and brain, and assess the resulting impact on viral replication and immune responses EX vivo. Aim 2 will characterize disease progression in ANKEF1-deficient mice by measuring viral titers, immune responses, and tissue damage in both the respiratory system and brain. Additionally, we will use single-cell RNA sequencing and spatial transcriptomics to identify key pathways associated with increased disease severity. Aim 3 will elucidate the molecular mechanisms by which ANKEF1 modulates ciliary functions and identify key interactions and regulatory networks that contribute to host defense against IAV infection. This research will provide new insights into the role of ciliary function in airway and brain defenses, potentially revealing therapeutic targets for severe influenza and other respiratory diseases.