American Society of Hematology

Figure 5

$Figure 5. Nuclear USP7 migrates to cytoplasm to inhibit TLR signal. Nuclear and cytoplasmic fractions of LPS-stimulated 293-TLR4 (A) and LPS and CpG ODN-stimulated THP1 (B) cells were collected at the indicated time points and immunoblotted with anti-USP7. Nucleolin and α-tubulin were blotted as nuclear and cytoplasmic fraction controls, respectively. (C) 293-TLR4 cells were transfected with nonsilencing shRNA or shRNA against A20, Cyld, or USP7 and knockdown of individual proteins was confirmed. Seventy-two hours later, TLR4 was stimulated with LPS for 6 hours before TNF-α and IL8 mRNA were assayed by qRT-PCR. (D) 293-TLR2/6 cells were transfected with empty vector (EV), full-length USP7 (USP7.FL), USP7.TD (aa's 1-210), USP7-ΔTD (aa's 210-1102), and USP7.C223S, and 24 hours later, TLR2/6 was stimulated with Pgn for 6 hours before TNF-α mRNA was assayed by qRT-PCR. Error bars represent SEM.$

Nuclear USP7 migrates to cytoplasm to inhibit TLR signal. Nuclear and cytoplasmic fractions of LPS-stimulated 293-TLR4 (A) and LPS and CpG ODN-stimulated THP1 (B) cells were collected at the indicated time points and immunoblotted with anti-USP7. Nucleolin and α-tubulin were blotted as nuclear and cytoplasmic fraction controls, respectively. (C) 293-TLR4 cells were transfected with nonsilencing shRNA or shRNA against A20, Cyld, or USP7 and knockdown of individual proteins was confirmed. Seventy-two hours later, TLR4 was stimulated with LPS for 6 hours before TNF-α and IL8 mRNA were assayed by qRT-PCR. (D) 293-TLR2/6 cells were transfected with empty vector (EV), full-length USP7 (USP7.FL), USP7.TD (aa's 1-210), USP7-ΔTD (aa's 210-1102), and USP7.C223S, and 24 hours later, TLR2/6 was stimulated with Pgn for 6 hours before TNF-α mRNA was assayed by qRT-PCR. Error bars represent SEM.

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