FIG. 6.
p16INK4A loss is responsible for conferring anchorage independence. (A) Pools of LTRas and anchorage-independent dnp53Ras Schwann clones that no longer expressed endogenous p16INK4A, expressing either wild-type rat p16INK4A or the empty vector pWZLhygro, were seeded in soft agar. The graph is representative of two independent experiments showing the average percentage of cells seeded that proliferated to form colonies in soft agar. The error bars represent the standard deviations of triplicate assays. The levels of p16INK4A as determined by immunoblotting are shown in the inset panel. (B) Populations of drug-selected Schwann cells, expressing dnp53 with the p16INK4A-insensitive mutant of cdk4 (cdk4R24C), antisense p16INK4A, or the N terminus of LT, together with Ras or the empty vector LXSN, were seeded in agarose. After 2 weeks the colonies were stained with neutral red prior to being counted and photographed.