Terminal differentiation and cellular senescence display common properties including irreversible growth arrest. To define the molecular and ultimately the biochemical basis of the complex physiological changes associated with terminal differentiation and senescence, an overlapping-pathway screen was used to identify genes displaying coordinated expression as a consequence of both processes. This approach involved screening of a subtracted cDNA library prepared from human melanoma cells induced to terminally differentiate by treatment with fibroblast IFN and mezerein with mRNA derived from senescent human progeria cells. overlapping-pathway screen ͉ terminal cell differentiation ͉ senescent phenotype ͉ interferon-inducible gene ͉ evolutionary conserved gene P lasticity of the transformed phenotype is suggested by the ability of differentiation-inducing agents to revert the cancerous properties of specific tumors (1-3). This attribute of tumor cells provides the basis for a potentially less toxic form of therapy, ''differentiation therapy.'' In metastatic human melanoma, a combination of IFN- and the protein kinase C activator mezerein (MEZ) produces irreversible growth arrest, a loss of tumorigenic competence, and terminal differentiation (1, 4). To define gene-expression changes associated with induction of terminal differentiation, a subtracted cDNA library enriched for genes associated with terminal differentiation was constructed (5). This construction was accomplished by subtracting control HO-1 human melanoma mRNAs from IFN- ϩ MEZ-treated HO-1 mRNAs, which were temporally collected over a 24-h period (5). This subtracted cDNA library then was screened by random isolation of phage colonies and Northern blotting, high-density cDNA microarray analysis, and reverse Northern screening followed by Northern blotting (5-7). These approaches have identified both unknown and known genes associated with tumor and normal growth control, cell-cycle regulation, IFN response, differentiation, and apoptosis (5-12). Four classes of melanoma differentiation-associated (mda) genes have been identified (5, 10).Terminal cell differentiation and cellular senescence are characterized by changes in cell morphology, lack of responsiveness to mitogenic stimulation, and irreversible growth arrest (1, 4, 13-18).Normal cells cultured in vitro lose their proliferative potential after a finite number of doublings in a process described as cellular senescence (13). Experiments in human diploid fibroblasts and additional cell types document an inverse correlation between replicative senescence and donor age and a direct relationship between replicative senescence and donor-species life span (13,19,20). In agreement with this relationship, cells from patients with premature aging syndromes such as Werner's syndrome and progeria achieve a quiescent state more rapidly than normal human fibroblasts (21). Although senescence is a time-dependent process, terminal differentiation can be induced in a variety of cell types by appropriate treatme...