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. 2022 Apr 29;13(4):414. doi: 10.1038/s41419-022-04801-0

Fig. 2. Midkine associates with AMPK subunits and its upstream regulating factors.

Fig. 2

a AMPK kinase subunit and LKB1 substrates were identified as MDK-associated proteins in HEK293A cells by tandem affinity purification–mass spectrometry. Bait MDK protein is marked in red. AMPKα is marked in blue. The right column of numbers represents unique peptide number/total peptide number. b, c MDK associates with LKB1, CAMKKβ, and AMPK subunits. The indicated constructs were expressed in the HEK293T cells for 24 h, and cell lysates were subjected to pull-down assays with S protein beads. d MDK was immunoprecipitated from HCCLM3 cells and subjected to Western blot analysis with antibodies against LKB1, Mo25a, STRADα, and MDK. e–f LKB1 immunoprecipitated from HCCLM3 (e) and HepG2 (f) cells and subjected to Western blot analysis with antibodies against MDK, Mo25a, STRADα, and LKB1. g–h HEK293T cells were cotransfected with MDK-HA and Flag-tagged LKB1 (g) or Myc-tagged Mo25a (h) and coimmunoprecipitated with HA primary antibody and subjected to Western blot analysis with antibodies against LKB1, HA, Mo25a and MDK.