The expression of FAC is regulated during cell cycle progression. HeLa cells were synchronized at G1 /S by double thymidine block and released into regular media. Cell aliquots (1 mL) from the indicated timepoints were washed, suspended in propidium iodide, and assayed by FACS analysis, as described in Materials and Methods. The percentage of cells in each phase of the cell cycle was determined by analyzing FACS data with the computer program CELLFIT (Becton Dickinson). Synchronized cells from each time interval were lysed, whole cell extracts were prepared, and cellular proteins were immunoprecipitated and immunoblotted with an antiserum against FAC (top) or immunoprecipitated with monoclonal against cdc2 followed by immunoblotting with anti–phosphotyrosine-cdc2 (middle) or anti-cdc2 (bottom). Immunoblots were scanned and analyzed by NIH Image. Values along the y-axis are in relative density units and are plotted against time in hours after release from G1 /S synchrony. Each plot is placed along side the corresponding immunoblot with symbols as follows: FAC (•); tyrosine phosphorylated cdc2 (▵); and total cdc2 (□).
