A PRE-START CHECKPOINT PREVENTING MITOSIS IN FISSION YEAST ACTS INDEPENDENTLY OF P34(CDC2) TYROSINE PHOSPHORYLATION

We have monitored the tyrosine (Y15) phosphorylated and dephosphorylated forms of p34(cdc2) from Schizosaccharomyces pombe as cells proceed through the cell cycle, Y15 is dephosphorylated in G(1) before start and becomes phosphorylated only after cells pass start and enter late G(1). This transition is associated with a switch from one checkpoint which restrains mitosis in pre-start G(1), by a mechanism independent from Y15 phosphorylation, to a second checkpoint acting post-start during late G(1) and S phase operating through Y15 phosphorylation, The pre-start checkpoint may act by preventing formation of the p34(cdc2)/p56(cdc13) complex, The complex between Y15-phosphorylated p34(cdc2) and p56(cdc13) accumulates during S phase and G(2), but the level generated is not solely dependent on the amount of p34(cdc2) and p56(cdc13) present in the cell, The extent of p56(cdc13) breakdown at the end of mitosis may be determined by the amount complexed with p34(cdc2). We have also shown that an insoluble form of p34(cdc2) is associated with the progression of the cell through late G(1) into S phase.