Figure 4.

ssRNAs from CVB3-Infected Cells Do Not Activate MDA5

(A) ssRNA fraction from mock- or CVB3-infected HeLa cells were transfected into MEFs of indicated genotypes, and IFN-β mRNA level at 2 hr.p.t. was measured by real-time qPCR. Data presented as mean ± SD.

(B) ssRNA fractions (ssPools) of mock- and CVB3-infected cells were treated with 10 ng/μl RNase A, 10 mU RNase III, or 10 mU DNase I at 37°C for 15 min. Resulting RNA samples equivalent to 500 ng starting material were transfected into approximately 200,000 MAVS^+/+ MEFs and IFN-β mRNA induction was determined by real-time qPCR at 8 hr.p.t. Data presented as mean ± SD.

(C) The same samples as in (B) were analyzed on agarose gel.

(D) CVB3 vRNA was isolated from pelleted viral particles. Indicated amounts of vRNA were transfected into RIG-I^−/− MEFs in the presence of CHX (10 μg/ml), and IFN-β mRNA induction was determined by real-time qPCR at 8 hr.p.t.. Data presented as mean ± SD.

(E) Indicated amounts of mock-treated or unlinkase-treated poliovirus vRNAs were transfected into RIG-I^−/− MEFs in the presence of CHX (10 μg/ml). Total RNA was extracted at 8 hr.p.t. and IFN-b mRNA induction was determined by real-time qPCR. Data presented as mean ± SD.

See also Figure S3.