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. 2014 Sep 12;33(22):2606–2622. doi: 10.15252/embj.201489244

Figure 2. The virulence effector OspF blocks formation of HP1γS83p in Shigella-infected cells.

Figure 2

  1. Shigella inhibits PMA-induced formation of HP1γS83p. HeLa cells were infected with the Shigella flexneri invasive strain (WT) and stimulated or not by PMA at the indicated times. Western blots were performed with anti-HP1γS83p, anti-HP1γ, and anti-actin antibodies.
  2. Pharmacological inhibition of the MAPK/MSK1 pathway blocks formation of HP1γS83p. HeLa cells were pretreated for 1 h with the MEK1 inhibitor U0126 or MSK1 inhibitor H89 and stimulated by PMA. Western blots were performed with anti-HP1γS83p, anti-HP1γ, anti-H3K9me2S10p, or anti-H3 antibodies.
  3. HeLa cells were infected with the Shigella flexneri invasive (WT) or the ospF-deficient (ospF) strains at the indicated time or stimulated by PMA (60 min). Western blots were performed with anti-HP1γS83p, anti-HP1γ, and anti-actin antibodies.
  4. Immunoprecipitation of endogenous HP1γ from HeLa cells overexpressing OspF. HeLa cells were transiently transfected with the indicated plasmids. Cellular extract were immunoprecipitated with mouse IgG as a negative control or HP1γ antibodies. Western blots were performed with OspF and HP1γ antibodies. The arrow indicates the OspF signal.
  5. Dephosphorylation at S83 in HP1γ immunoprecipitates containing OspF. HeLa cells were transiently transfected with the indicated plasmids and cellular extract immunoprecipitated with HP1γ antibody in the absence or the presence of DNase I. Western blots were performed with anti-HP1γS83p, anti-HP1γ, anti-OspF, or anti-ERK antibodies.

Source data are available online for this figure.