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. 2022 Sep 29;1(5):100057. doi: 10.1016/j.cellin.2022.100057

Fig. 7.

Fig. 7

Trim25 disrupts the stability of RABV-P. (A) 293T cells were co-transfected with FLAG-Trim25 (at the indicated volumes), HA-CVS-P, and HA-GFP for 48 h, and cell lysates were analyzed by Western blotting. (B) 293T cells were transfected with pCAGGS, HA-CVS-P, FLAG-Trim25 with pCAGGS, or FLAG-Trim25 with HA-CVS-P for 36 h. Then, cellular RNA was extracted, and CVS-P mRNA levels were measured. (C) 293T cells were co-transfected with FLAG-Trim25, HA-CVS-P, and HA-GFP for 48 h, and 10 μM CHX was added to the culture medium at 0, 3, 6, and 9 h before cell lysis. The cell lysates were analyzed by Western blotting. (D) 293T cells were co-transfected with FLAG-Trim25 and HA-CVS-P for 48 h and treated with 10 μM MG132 10 h before cell lysis. The cell lysates were analyzed by Western blotting. (E, F) 293T cells were co-transfected with HA-74-297-P, HA-74-297-P with FLAG-Trim25 (E) or FLAG-E3-mut-Trim25 (F), HA-CVS-P, and HA-CVS-P with FLAG-Trim25 (E) or FLAG-E3-mut-Trim25 (G) for 48 h. Then, cell lysates were analyzed by Western blotting. (G, H) 293T cells were co-transfected with HA-72m-P, HA-72m-P with FLAG-Trim25 (G) or FLAG-E3-mut-Trim25 (H), HA-CVS-P, and HA- CVS-P with FLAG-Trim25 (G) or FLAG-E3-mut-Trim25 (H) for 48 h. Then, cell lysates were analyzed by Western blotting. Western blotting data are representative of at least two independent experiments. See also Fig. S5.