FIGURE 2.
p62 interacts with IRS-1 through its SH2 domain. A, domain structures of full-length p62 and p62 mutants (ΔUBA, ΔN-term, and ΔSH2) were used to map the interaction with IRS-1. N, N terminus; AID, acidic interaction domain; LIR, LC3-interacting region; C, C terminus. B, CHO/IR cells were transfected with IRS-1 and wild-type Myc-tagged p62, ΔUBA, ΔN-term, or HA-p62ΔSH2 domain. The cells were stimulated with insulin (100 nm) for 15 min and lysed in Triton lysis buffer. The cell lysates were immunoprecipitated (IP) with anti-V5 followed by Western blotting with anti-IRS-1 and anti-Myc/HA. The lysates were also blotted with anti-Myc/HA to verify the expression of the p62 constructs. Experiments were replicated three times with similar results.