Commitment Point during G₀→G₁ That Controls Entry into the Cell Cycle

Abstract

Initiation of T-lymphocyte-mediated immune responses involves two cellular processes: entry into the cell cycle (G 0 3G 1 ) for clonal proliferation and coordinated changes in surface and secreted molecules that mediate effector functions. However, a point during G 0 3G 1 beyond which T cells are committed to enter the cell cycle has not been defined. We define here a G 0 3G 1 commitment point that occurs 3 to 5 h after CD3 and CD28 stimulation of human CD4 or CD8 T cells. Transition through this point requires cdk6/4-cyclin D, since inhibition with INK4A during the first 3 to 5 h prevents cell cycle entry and maintains both naive and memory T cells in G 0 . Transition through the G 0 3G 1 commitment point is also necessary for T cells to increase in size, i.e., to enter the cellular growth cycle. However, transition through this point is not required for the induction of effector functions. These can be initiated while cells are maintained in G 0 with TAT-p16 INK4A . We have termed this quiescent, activated state G 0(A) . Our data provide proof of the principle that entry of T cells into the cell cycle and cellular growth cycles are coupled at the G 0 3G 1 commitment point but that these processes can be uncoupled from the early expression of molecules of effector functions.Many different cell types in the human body are in a quiescent (G 0 ) state. In order to proliferate, such cells must first enter G 1 before progressing through the cell cycle and undergoing division to produce two daughter cells. Progression through the cell cycle is dependent on the integration of mitogenic signals, assessment of cell size, and DNA integrity. There is a point late in G 1 beyond which cells no longer need mitogenic stimulation to enter S phase and proliferate, and this is known as the restriction point (42). Commitment to enter the cell cycle from G 0 is similarly important, but a point that controls the G 0 3G 1 transition and commitment to enter the cell cycle has not been defined.T lymphocytes are one of the few available primary cell models commonly used for studying cell cycle entry and progression. The majority of primary peripheral blood T lymphocytes rest in a true G 0 state, and these can be activated through their T-cell receptor and CD28 costimulation to undergo both cell cycle progression and functional activation. T-lymphocytemediated immune responses involve two simultaneous processes. The first is entry into the cell cycle to expand naive T-lymphocyte clones. As they progress through the cell cycle, cells must increase in size in order that their cellular contents are maintained with each round of cell division. This process has been termed the cellular growth cycle (62). The second process is a change in surface and secreted molecules that mediate effector functions (19), giving rise to differentiation of T lymphocytes into effector or memory phenotype. Activation and clonal expansion of T cells occur normally upon interaction with an antigen-presenting cell. A defined program is thus initiated (55, 56) th...

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