Threshold expression of cyclin E but not D type cyclins characterizes normal and tumour cells entering S phase.

Abstract

Complexes of cyclin-dependent kinases (cdk) and their partner cyclins drive the cell through the cell cycle, each such complex phosphorylating a distinct set of proteins at a particular check-point or phase of the cycle. Immunocytochemical detection of cyclins combined with measurement of cellular DNA content by flow cytometry makes it possible to relate expression of each of these proteins with the actual cell cycle position, without the necessity of cell synchronization. In the present study, we have investigated expression of E and D type cyclins in G1 cells and in cells entering S phase, in eight different human hematopoietic and solid tumour cell lines (two leukaemias, a lymphoma, three breast carcinomas, a colon carcinoma and a bladder transitional cell carcinoma) during their exponential phase of growth, as well as in normal mitogen stimulated lymphocytes. In all the cell types studied, the average level of D type cyclin expression was invariable throughout the cell cycle. A great intercellular variability, in particular of the G1 cell subpopulations, and the presence of a large fraction of G1, S and G2 + M cells that were cyclin D negative (20-40% in tumour cell lines and about 80% among lymphocytes), were other characteristic features of D type cyclin expression. In contrast to D type cyclins, the expression of cyclin E was discontinuous during the cycle, peaking at the time of cell entrance to S. Also, a well defined threshold in expression of cyclin E characterized cells that were entering S phase, and virtually no cyclin E negative cells were seen during the early portion of S phase. The data indicate that while cell entrance to S phase is unrelated to expression of D type cyclins (at the time of entrance), accumulation of cyclin E up to critical level is a prerequisite for initiation of DNA replication. The great intercellular variability in expression of D type cyclins and their invariant average level across the cell cycle suggest that these cyclins, in addition to their acknowledged function in promoting cell progression through mid- to late-G1 may have other role(s), related or unrelated to the cell cycle progression. The presence of a large number of D type cyclin negative cells in all phases of the cycle suggests that during exponential growth the cells may not express this protein and yet may traverse the cycle, including G1 phase.