Skip to main content
. Author manuscript; available in PMC: 2016 Apr 21.
Published in final edited form as: J Cell Physiol. 2015 May;230(5):1075–1085. doi: 10.1002/jcp.24839

FIGURE 6. Influence of MATα2 and MATβ phospho-region mutations on HSC activation.

FIGURE 6

A. WT and phospho-mutant vectors of MAT2α2 (DDK-tagged α2-P1, P2 and P3) were expressed in HSCs as described under Materials and Methods and total cellular extracts were immunoblotted with MATα2-DDK tag, endogenous MATα2, α-SMA, actin and α-tubulin antibodies. Interaction of MATα2-DDK and its mutants with exogenous MATβ-HA protein were estimated by co-immunoprecipitation (IP: β-HA blots). Densitometric ratios normalized to actin are represented as fold over EV for α-SMA and fold over WT MATα2 for mutant proteins. Results are mean ± S.E. from 6 experiments. *p<0.05, **p<0.005, †p<0.001, vs. EV or α2-DDK over-expression. B. WT and phospho-mutant vectors of MATβ (HA-tagged β-P1 and P2) were over-expressed in HSCs and subjected to phos-tag™ analysis as described in Materials and Methods. Results are representative of three analysis. C. WT and phospho-mutant vectors of MATβ (HA-tagged β-P1 and P2) were expressed in HSCs as described under Materials and Methods and total cellular extracts were immunoblotted with MATβ-HA tag, endogenous MATβ, α-SMA, actin and α-tubulin antibodies. Interaction of MAT β-HA and its mutants with exogenous MATα2-DDK protein were estimated by co-immunoprecipitation (IP: α2-DDK blots). Densitometric ratios normalized to α-tubulin are represented as fold over EV for α-SMA and fold over WT MATβ for mutant proteins. Results are mean±S.E. from 6 experiments. **p<0.005, *p<0.05, ¥p<0.01 vs. EV or β-HA over-expression. D. RNA from cells over-expressing MAT2A-DDK (left panel) or MAT2B-HA (right panel) vectors and their mutants was measured by relative quantitative RT-PCR and expressed as fold over EV. Results are representative of three independent experiments. *p<0.05, **p<0.005 vs. EV.