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. Author manuscript; available in PMC: 2015 Aug 25.
Published in final edited form as: Dev Cell. 2014 Aug 7;30(4):394–409. doi: 10.1016/j.devcel.2014.06.013

Figure 5. TRIM5α interacts with activated Beclin 1 and assembles ULK-1 with Beclin 1.

Figure 5

(A) Domain organization of TRIM5α and deletion constructs used in mapping experiments in B. (B) Co-immunoprecipitation analysis of full-length or deletion mutants of GFP-TRIM5α with FLAG-tagged Beclin 1. (C) Co-immunoprecipitation analysis of interactions between indicated TRIMs (as GFP fusions) and FLAG-Beclin 1 in 293T cell extracts. (D) Bcl-2-Beclin 1 complexes assessed by co-immunoprecipitation from control cells or cells expressing HA-HuTRIM5α or HA-RhTRIM5α. (E) Abundance of TAB2-Beclin 1 complexes assessed by co-immunoprecipitation from 293T expressing GFP-TRIM5α or GFP alone. Transfected cells were subjected to TAB2 immunoprecipitation and intensities of Beclin 1 bands in the precipitates normalized to TAB2 in the same samples. Differences in the normalized values (set at 100% for GFP-expressing cells) were assessed between cells expressing GFP or GFP-TRIM5α. Data, means ± SE, n = 3 experiments, *, P < 0.05 (t test). (F) Assessment of TRIM5α effects on ULK1 presence in Beclin 1 complexes. 293T cells were transiently transfected with Myc-ULK1, FLAG-Beclin 1, and either GFP-TRIM5α or GFP alone. Lysates were immunoprecipitated with anti-FLAG, and immunoblots probed as indicated. (G) Lysates from 293T cells expressing GFP or the indicated GFP-TRIMs, Myc-ULK1, and FLAG-Beclin 1 were treated as in F. See also Figure S5.