Oncogene-induced senescence is a tumor-suppressive defense mechanism triggered upon activation of certain oncogenes in normal cells. Recently, the senescence response to oncogene activation has been shown to act as a bona fide barrier to cancer development in vivo. Multiple previous studies have implicated the importance of the p38 MAPK pathway in oncogene-induced senescence. However, the contribution of each of the four p38 isoforms (encoded by different genes) to senescence induction is unclear. In the current study, we demonstrated that p38␣ and p38␥, but not p38␤, play an essential role in oncogenic ras-induced senescence. Both p38␣ and p38␥ are expressed in primary human fibroblasts and are activated upon transduction of oncogenic ras. Small hairpin RNA-mediated silencing of p38␣ or p38␥ expression abrogated ras-induced senescence, whereas constitutive activation of p38␣ and p38␥ caused premature senescence. Furthermore, upon activation by oncogenic ras, p38␥ stimulated the transcriptional activity of p53 by phosphorylating p53 at Ser 33 , suggesting that the ability of p38␥ to mediate senescence is at least partly achieved through p53. However, p38␣ contributed to ras-inducted senescence via a p53-indepdendent mechanism in cells by mediating ras-induced expression of p16 INK4A , another key senescence effector. These findings have identified p38␣ and p38␥ as essential components of the signaling pathway that regulates the tumor-suppressing senescence response, providing insights into the molecular mechanisms underlying the differential involvement of the p38 isoforms in senescence induction.The ras proto-oncogenes encode small GTP-binding proteins that transduce growth signals from cell surface (1-3). Aberrant activation of ras is a crucial step in tumor formation. Constitutive activation of ras genes, either through point mutations or overexpression, is associated with a wide variety of human tumors at high frequency and contributes to the initiation and maintenance of multiple tumorigenic phenotypes in these cancers (4 -11). However, in early-passage primary human and rodent cells, activated ras causes a permanent proliferative arrest known as premature senescence, because of its phenotypic similarities to replicative senescence observed in late-passage cells (12). Other oncogenes, such as E2F1 and raf, or inactivation of certain tumor suppressor genes, also induce senescence in normal human cells (13-15). The existence of the premature senescence response to oncogene activation implies that like apoptosis, oncogene-induced senescence serves as an anti-tumorigenic defense mechanism. Indeed, it has been well documented that cellular transformation by ras requires cooperation from immortalizing oncogenes that overcome the senescence response, such as those inactivating p53 (8,16,17). Recent studies have also demonstrated that senescent cells can be detected in early-stage premalignant lesions of lung, pancreas, skin, and prostate in both human cancer patients and mouse tumor models and that disruption of s...

c Oncogene-induced senescence is a stable proliferative arrest that serves as a tumor-suppressing defense mechanism. p38 mitogen-activated protein kinase (MAPK) has been implicated in oncogene-induced senescence and tumor suppression. However, the specific role of each of the four p38 isoforms in oncogene-induced senescence is not fully understood. Here, we demonstrate that p38␦ mediates oncogene-induced senescence through a p53-and p16INK4A -independent mechanism. Instead, evidence suggests a link between p38␦ and the DNA damage pathways. Moreover, we have discovered a novel mechanism that enhances the expression of p38␦ during senescence. In this mechanism, oncogenic ras induces the Raf-1-MEK-extracellular signal-regulated kinase (ERK) pathway, which, in turn, activates the AP-1 and Ets transcription factors that are bound to the p38␦ promoter, leading to increased transcription of p38␦. These findings indicate that induction of the prosenescent function of p38␦ by oncogenic ras is achieved through 2 mechanisms, transcriptional activation by the Raf-1-MEK-ERK-AP-1/Ets pathway, which increases the cellular concentration of the p38␦ protein, and posttranslational modification by MKK3/6, which stimulates the enzymatic activity of p38␦. In addition, these studies identify the AP-1 and Ets transcription factors as novel signaling components in the senescence-inducing pathway.A lthough aberrant activation of Ras is associated with human tumors, activated ras in early-passage primary human and rodent cells causes permanent growth arrest known as oncogeneinduced senescence (OIS) (1-4). Like apoptosis, OIS is a tumorsuppressing defense mechanism, the disruption of which leads to tumorigenesis (5-10).Multiple signaling intermediates have been identified that play critical roles in the pathways mediating oncogene-induced senescence. The ability of ras to induce senescence depends on activation of the Raf-MEK-extracellular signal-regulated kinase (ERK) mitogen-activated protein kinase (MAPK) pathway (4, 11) and is accompanied by upregulation of p16 INK4A , p53, p14/p19 ARF , and p21 WAF1 (3, 12) and silencing of E2F target genes (13). We previously showed that ras-induced senescence relies on activation of p38, a MAPK previously identified as a major mediator of inflammation and stress responses (14). p38 and its upstream MAPK kinases MKK3 and MKK6 are activated by oncogenic ras as a result of persistent MEK/ERK activation in senescent cells. Constitutive activation of p38 causes premature senescence, whereas inhibition of p38 prevents ras-induced senescence (14). Consistent with the important role of p38 in oncogene-induced senescence and tumor suppression, targeted deletion of p38␣ or PRAK, a downstream substrate kinase of p38, accelerates cancer development in mouse models (10,15,16).p38 MAK has four mammalian isoforms, ␣, ␤, ␦, and ␥, which are encoded by different genes and differ in tissue-specific expression, substrate spectrum, and affinity for upstream MAPK activators (17-23). Our previous data indicated that p38...