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. 2020 Sep 15;11:581011. doi: 10.3389/fphar.2020.581011

Figure 4.

Figure 4

HCK interacts with NLRP3 independent of its PYD domain. (A) HEK293T cells were transfected with MYC-HCK (WT) and FLAG-NLRP3, and the interaction between HCK and NLRP3 was analyzed using immunoprecipitation (IP) with anti-FLAG antibody and immunoblotting (IB) with the anti-FLAG and anti-MYC antibodies. (B) HEK293T cells were transfected with MYC-HCK (WT) and FLAG-NLRP3 or its truncated forms (FLAG-PYD, FLAG-NBD, or FLAG-LRRs, followed by immunoprecipitation (IP) with anti-FLAG and immunoblotting (IB) with anti-FLAG or anti-MYC antibodies. (C) LPS-primed PMs were treated with A419259 and then stimulated with 5 μM nigericin for 30 min, followed by immunoprecipitation (IP) with HCK antibody and immunoblotting (IB) with antibodies against HCK, phosphorylated tyrosine, NLRP3, ASC or β-tubulin. (D) The normalized levels of HCK phosphorylation were quantified. Data from (A–C) are representative of at least three independent experiments. Data show means ± SEM, *P > 0.05, **P < 0.01, Student’s t-test.