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. 2018 Dec 4;29(2):136–150. doi: 10.1038/s41422-018-0120-9

Fig. 4.

Fig. 4

Ubiquitinated Rheb promotes its interaction with TSC. a Co-IP assay was performed to analyze the binding of TSC2 to different forms of Rheb mutants (active form: S16H; inactive form: D60V, D60K, S20N). b TSC2 purified from HEK293T cells showed stronger binding affinity to the ubiquitinated Rheb. c Rheb-K8R mutant showed weaker binding affinity to TSC2 as compared with Rheb-WT. d, e RNF152 Knockdown (d) or knockout (e) reduced the binding bewteen Rheb and TSC2. The knockdown efficiency was detected by RT-PCR in Supplementary information, Fig. S2e. RNF152-KO MEF cells were analyzed via genomic DNA PCR in Supplementary information, Fig. S3s. fh USP4 knockdown (f), knockout (g) or inactivation of USP4 by Vialinin A (h) promoted the binding of Rheb to TSC2