Fig. 7

Overexpression/inhibition of GACAT2 affects the expression, activity, allosteric regulation, post-translational modification and translocation of PKM1/2. a–d Relative protein expression levels of PKM1/2, PKM1 and PKM2 in PDLSCs (with overexpression or inhibition of GACAT2) determined by Western blots and semiquantitative analysis of their expression levels (normalized to β-actin) in terms of relative gray density. The cells were transfected with ov-NC or ov-GACAT2 and incubated in medium with both inflammatory cytokines and EMD (inflammatory environment) (a and b) or were transfected with si-NC or si-GACAT2 and incubated in medium with the cementoblastic inducer EMD (noninflammatory environment) (c and d). e–h PK activity and production of pyruvate in PDLSCs (with the overexpression or inhibition of GACAT2). The cells were transfected with ov-NC or ov-GACAT2 and incubated in an inflammatory environment (e and f) or were transfected with si-NC or si-GACAT2 and incubated in a noninflammatory environment (g and h). i–l Tetramer, dimer and monomer levels of PKM2 in PDLSCs (with overexpression or inhibition of GACAT2) determined by western blots (i and k) and semiquantitative analysis of their expression levels (normalized to β-actin) in terms of relative gray density (j and l). The cells were transfected with ov-NC or ov-GACAT2 and incubated in an inflammatory environment (i and j) or were transfected with si-NC or si-GACAT2 and incubated in a noninflammatory environment (k and l). m–p p-PKM2 in PDLSCs (with overexpression or inhibition of GACAT2) was determined by western blots and semiquantitative analysis of their expression levels (normalized to β-actin) in terms of relative gray density. The cells were transfected with ov-NC or ov-GACAT2 and incubated in an inflammatory environment (m and n) or were transfected with si-NC or si-GACAT2 and incubated in a noninflammatory environment (o and p). q Immunofluorescence staining for mitochondria (MitoTracker, red), PKM2 (green) and DAPI (blue) in the PDLSCs transfected with ov-NC or ov-GACAT2 and incubated in an inflammatory environment (scale bar: 50 μm). r and s Relative protein expression levels of mitochondrial PKM2 in PDLSCs (with overexpression of GACAT2) determined by Western blots (r) and semiquantitative analysis of their expression levels (normalized to COXIV) in terms of relative gray density (s). The data are shown as the mean ± SD for n from 3 to 6; *P < 0.05, **P < 0.01 and ***P < 0.001 indicate significant differences between the indicated columns