Loss of p53 induces cell proliferation via Ras-independent activation of the Raf/Mek/Erk signaling pathway

Abstract

The Ras family of small GTPases constitutes a central node in the transmission of mitogenic stimuli to the cell cycle machinery. The ultimate receptor of these mitogenic signals is the retinoblastoma (Rb) family of pocket proteins, whose inactivation is a required step to license cell proliferation. However, little is known regarding the molecular events that connect Ras signaling with the cell cycle. Here, we provide genetic evidence to illustrate that the p53/ p21 Cdk-interacting protein 1 (Cip1)/Rb axis is an essential component of the Ras signaling pathway. Indeed, knockdown of p53, p21Cip1, or Rb restores proliferative properties in cells arrested by ablation of the three Ras loci, H-, N-and K-Ras. Ras signaling selectively inactivates p53-mediated induction of p21Cip1 expression by inhibiting acetylation of specific lysine residues in the p53 DNA binding domain. Proliferation of cells lacking both Ras proteins and p53 can be prevented by reexpression of the human p53 ortholog, provided that it retains an active DNA binding domain and an intact lysine residue at position 164. These results unveil a previously unidentified role for p53 in preventing cell proliferation under unfavorable mitogenic conditions. Moreover, we provide evidence that cells lacking Ras and p53 proteins owe their proliferative properties to the unexpected retroactivation of the Raf/Mek/Erk cascade by a Ras-independent mechanism.T he RAS genes have been extensively studied due to their key role in mediating mitogenic signaling as well as their high prevalence in human cancers, including those cancers with poor survival rates, such as lung adenocarcinoma, colorectal carcinoma, and pancreatic ductal adenocarcinoma (1, 2). However, the mechanisms by which Ras proteins mediate mitogenic signaling in either normal or tumor cells remain obscure, especially beyond activation of the Raf/Mek/Erk cascade. Recent genetic studies have underscored the relevance of Ras proteins in cellular homeostasis by demonstrating that cells lacking the three Ras loci, H-Ras, N-Ras, and K-Ras (Rasless cells), are completely unable to proliferate (3, 4). Indeed, systemic ablation of these loci in adult mice causes rapid deterioration of multiple tissues, leading to their death in a few days.GTP-loaded Ras proteins promote activation of various downstream signal transduction pathways, mainly the Raf/Mek/Erk kinase cascade, the PI3K/Akt route, and the Ral guanine dissociation stimulator (RalGDS) pathway (1). Activation of other pathways, such as those pathways driven by the Rac family of small G proteins and phospholipase C, has also been illustrated (1). However, genetic interrogation of the pathways essential for cell proliferation has illustrated that only constitutive activation of the Raf, Mek, or Erk kinase can bypass the requirement for Ras proteins to sustain cell division, at least in vitro (3, 4). Indeed, constitutive activation of the PI3K/Akt and RalGDS pathways was incapable of inducing cell proliferation in the absence of Ras proteins. In agreement wit...

PDCD5 regulates cell proliferation, cell cycle progression and apoptosi

“…Loss of p53 was recently found to induce cell proliferation via Ras-independent activation of the Raf/Mek/Erk signaling pathway (70). In line with these findings, overexpression of p53 is known to inhibit cell proliferation (71,72).…”

Ras and p53: An unsuspected liaison

“…This screen unambiguously identified the Trp53/Cdkn1a (p53/p21Cip1) tumor suppressor axis as an essential mediator of Ras mitogenic signaling. 7 Indeed, efficient knockdown of either p53 or p21Cip1 expression fully restored the proliferative properties of Rasless cells (Fig. 1B).…”

“…Further studies revealed that loss of Ras proteins caused widespread transcriptional activation of p53 through a mechanism involving acetylation of 2 specific residues in its DNA binding domain. 7 Surprisingly, phosphorylation and/or protein stabilization of p53 was not required. These genetic studies place the p53/p21 axis at the center of the mitogenic pathway that connects Ras signaling with the cell cycle (Fig.…”

Genetic Validation of Cell Proliferation via Ras-Independent Activation of the Raf/Mek/Erk Pathway

“…We have recently shown that absence of the p53/ p21Cip1/Rb axis indeed causes such a Ras-independent Raf/Mek/ Erk activation and allows proliferation of Rasless MEFs [ 9 ]. No rescue in the absence of p53, p21Cip1, or Rb was observed in Rafl ess, Mekless, or Erkless cells, indicating that, in contrast to Ras proteins, the Raf, Mek, or Erk kinases were absolutely essential for cell proliferation.…”

“…Interestingly, addition of a membrane-attachment domain to the Raf kinases was suffi cient to bypass the requirement for Ras proteins, thus suggesting that the major function of Ras proteins in mitogenic signaling is to recruit Raf kinases to the membrane, where activation of their kinase activity must occur independently of Ras [ 8 ]. Likewise, genetic elimination of the three Raf isoforms (A-Raf, B-Raf, c-Raf), both Mek (Mek1 and 2) or both Erk isoforms (Erk1 and 2) caused a cell cycle arrest phenotype indistinguishable from that of Rasless cells [ 9 ]. Taken together, these observations indicate that activation of the Raf/Mek/Erk pathway by Ras proteins is both necessary and suffi cient to sustain cell proliferation.…”