Fig 2.

Inhibition of translation disrupts P-body formation, and blocks in the catalytic steps of decapping or 5′-to-3′ degradation lead to increased P-body formation. (A) Fluorescence micrographs of strains expressing Dcp2-tdTomato or GFP fusion proteins of various P-body components grown at 30°C (control) or after exposure to 100 μg/ml cycloheximide (CHX) for 15 min or in the exo2 mutant genetic background are shown. Scale bar, 5 μm. (B) Fluorescence micrographs of dcp2 shutoff strains expressing GFP fusion proteins of various P-body components under derepression conditions (nmt41 promoter on) or after depletion of Dcp2 by addition of thiamine to the medium for 16 h are shown (left panel). Total cell lysates separated by SDS-PAGE were subjected to Western blotting using anti-HA to reveal Dcp2 protein levels (upper right panel). Antibodies against α-tubulin were used as a loading control. Growth curves of dcp2 shutoff strains after depletion of Dcp2 by the addition of thiamine to medium are shown (lower right panel).