Figure 6.
Forced Altered Localization of TRIM5α by Overexpression of TRIM5α or Inhibition of SUMOylation Reduces Type I IFN Induction following Retroviral Infection
Human DCs were DOTAP transfected with pLPCX TRIM5α-HA (T5) or empty pLPCX (EV) at 1 μg DNA/0.1 × 106 DCs, or left untransfected (day 0) ± 100 μM GA or 100 μM TDF (day 1) and infected with indicated retroviruses at MOI 5 (day 2).
(A) TRIM5α mRNA expression normalized for housekeeping transcript RPL13A (60S ribosomal protein L13a). NI EV cells were arbitrarily set to 1.
(B) Infection efficiency was assessed by qPCR of eGFP/eYFP DNA in DCs extracts at 10 hpi. eGFP/eYFP signal was normalized for housekeeping gene GAPDH. NT or EV were arbitrarily set to 100% for each infection. Signal was undetectable for non-infected samples.
(C–E) IFNα1 (C), IFNβ (D), and IL6 (E) mRNA expression levels were assessed by reverse transcription qPCR at 24 hpi, normalized for housekeeping transcript β2M. NI NT cells were arbitrarily set to 1. All panels are representative of at least three independent experiments on different donors.
See also Figure S4.