Fig. 4.

Induction of an ectopic kinetochore causes cellular transformation. a 3T3 cells with or without an integrated lacO array and expressing GFP-LacI or GFP-CENP-T-ΔC-LacI were grown for 4 days in the presence or absence of IPTG as indicated. Cells were then plated in triplicate in soft agarose in the presence of IPTG. After 4 weeks of growth, 200 cells were counted per replicate, and the percentage of cells forming colonies was scored. Clonal cell lines expressing (high) or (low) levels of the fusion protein were tested for each condition as indicated. Primary mouse embryonic fibroblasts (MEFs) and HeLa cells were used as negative and positive controls, respectively. Asterisk indicates significant difference as determined by T test, P < 0.001. b 3T3-lacO cells expressing GFP-LacI or GFP-CENP-T-ΔC-LacI were grown for 14 days in the presence or absence of IPTG as indicated. Cells were then seeded into the upper chamber of a basement membrane-coated Boyden’s chamber and migration across the membrane was assayed at the indicated time points by staining with crystal violet. N = 20 fields. Error bars show standard deviation. Right hand panel shows images of lower membrane staining from a representative experiment