FIGURE 2.

SHMT1 protein levels change with the cell cycle. HeLa cells were cell cycle-blocked by 24-h treatment with lovastatin (30 μm) for G₁ phase, hydroxyurea (1 mm) for S phase, and nocodazole (100 ng/ml) for G₂/M phase. A, SHMT1 protein levels were determined by immunoblotting of 20 μg of protein extract. B, SHMT1 mRNA levels were quantified from whole cell extract. C, Ub-SHMT1 levels were quantified by immunoprecipitation (IP) of ubiquitinated substrates from whole cell extracts (20 μg) followed by SHMT1 immunoblotting (IB). All experiments were repeated at least three times with similar results.