Recurrent rhinovirus infections in a child with inherited MDA5 deficiency

Lamborn, Ian T.; Jing, Huie; Zhang, Yu; Drutman, Scott B.; Abbott, Jordan K.; Munir, Shirin; Bade, Sangeeta; Murdock, Heardley M.; Santos, Celia P.; Brock, Linda G.; Masutani, Evan; Fordjour, Emmanuel Y.; McElwee, Joshua J.; Hughes, Jason D.; Nichols, Dave P.; Belkadi, Aziz; Oler, Andrew J.; Happel, Corinne S.; Matthews, Helen F.; Abel, Laurent; Collins, Peter L.; Subbarao, Kanta; Gelfand, Erwin W.; Ciancanelli, Michael J.; Casanova, Jean-Laurent; Su, Helen C.

The patient's infection history suggested that MDA5 might function as a general sensor of viruses infecting the respiratory tract in humans, including HRV, RSV, and influenza virus. Alternatively, HRV, as a member of the picornavirus family characterized by a single positive sense strand RNA genome, would be more likely to have extended stretches of RNA double helices during the generation of viral replicative intermediates that might bind effectively to MDA5 (Palmenberg et al., 2009) . To test innate immune responses in respiratory epithelium, we infected a transformed respiratory epithelial cell line A549, in which genes of the RLR pathway were silenced by transient transfection of siRNA. Silencing of MDA5, RIG-I, or MAVS expression increased both HRV transcripts and production of infectious virus (Fig. 4, and Fig. S1, A and B) . The requirement for MDA5 or MAVS in optimal virus control occurred not only for HRV-B (Fig. 4 , A-F; and Fig. S1, A and B), but also for HRV-A in human primary fibroblasts (Fig. 4, G and H; and Fig. S1 C) . In A549 cells, HRV minimally induced low levels of IFN-regulated transcripts, which were decreased upon MDA5 silencing, consistent with further impairment in virus recognition and antiviral responses (Fig. 5, A and B ). Both IFN-β (IFNB1) Equivalent lysates from ∼30,000 cells were run across lanes. **, P < 0.01; ***, P < 0.001; ****, P < 0.0001, by one-way ANO VA. and IFN-λ (IFNL3, IL-28) transcripts were decreased upon MDA5 or MAVS silencing in primary human fibroblasts infected with HRV ( Fig. 5 , C-E). Increased HRV replication was also observed in primary respiratory nasal epithelial cells from the patient as compared with healthy controls or her parents ( Fig. 6 A and Fig. S1 D) . Furthermore, increased HRV replication was observed in SV40-transformed fibroblasts in which CRI SPR/Cas9 genome editing had been used to generate single-cell clones either hemizygous for the patient's mutant IFIH1 allele or completely lacking both IFIH1 alleles (Fig. 6 B and Fig. S1 E) . Importantly, transduction with wild-type but not mutant K365E MDA5 in A549 cells improved control of HRV replication (Fig. 6 , C and D). Thus, these data supported a role for MDA5 in the control of HRV infection in humans and for the patient's recurrent HRV infections.