Cell cycle regulation in acinar cell death

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Salivary gland hypofunction following chemotherapeutic regimens or head/neck irradiation presents a major oral health problem. Loss of salivary gland acinar cells under these conditions appears to be through apoptosis, a genetically programmed, energy requiring multi-step process. One of the mechanisms through which apoptotic stimuli might mediate cell death is through altered cell cycle regulation. Cell death in response to diverse stimuli including growth factor withdrawal, activation of 'death' receptors or DNA damage may be accompanied by ectopic activation of cyclin dependent kinases (cdks) which may lead to inappropriate cell cycle re-entry or progression in diverse types of cells, including secretory epithelial cells. Over-expression of cyclin dependent kinase inhibitors (CKIs), proteins that inhibit cdks and therefore cell cycleprogression, can protect cells from apoptosis associated with different stresses. It is our hypothesis that inappropriate activation of cyclin dependent kinases is an integral part of the apoptotic pathway in acinar cells, and that cyclin dependent kinase inhibitors can attenuate apoptosis through modulating cdk activity. Our preliminary data suggest that suggest that DNA increases the kinase activity of specific cyclin/cdk complexes in immortalized and in primary acinar cells. 'Ectopic' activation of cdks is an early event in acinar cell death, occurring at the same time as caspase-3 activation. Increased cyclin/cdk kinase activity also correlates with loss of expression of the CKI p27/kip1. In this proposal, we will examine the role of DNA damage-induced cdk activation in animal cell apoptosis. We will determine whether 'ectopic' cdk activation precedes or requires caspase--3 activation in primary cultures of acinar cells. We will resolve whether cdk activation is necessary for acinar cell death, and we will ascertain whether cdks phosphorylate normal cell cycle substrates or perhaps act on novel substrates during apoptosis. We will explore the ability of the CKIs/p27/kip1 and p21/cip1 to modulate DNA-damage induced apoptosis through cdk activity. In addition, we will verify the protective functions of either p21/cip1 or p27/kip1 in CKI null cells. Activation of the tumor suppressor 53 mediates DNA damage induced apoptosis through induction of p21/cip1 gene transcription in many types of cell. Therefore, we will examine the role of the tumor suppressor in acinar cell death by testing the effects of DNA damage and CKI expression in p53 null cells.