FIG. 9.

Association of eIF6 with 60S ribosomal subunits in yeast cells. An exponentially growing culture of KSY603 was harvested, the cells were washed and lysed, and the cell lysate was subjected to 5 to 30% sucrose gradient centrifugation to separate polysomes, free 80S ribosomes, and ribosomal subunits as described in the legend to Fig. 5 and Materials and Methods. Fractions from the gradient were collected, and proteins were precipitated with 10% trichloroacetic acid, analyzed by SDS-PAGE, and immunoblotted with either anti-L3 antibodies or anti-HA antibodies as probes. Fractions containing 40S and 60S subunits, 80S monosomes, and polysomes are indicated. Protein L3 was used as a marker for 60S subunit sedimentation.