p53 regulation by post-translational modification and nuclear retention in response to diverse stresses Gretchen S Jimenez 1 , Shireen H Khan 1,2 , Jayne M Stommel 1,2 and Georey M Wahl* ,1 1 Gene Expression Laboratory, The Salk Institute, 10010 N. Torrey Pines Road, La Jolla, California, CA 92037, USA; 2 Department of Biology, University of California at San Diego, La Jolla, California, CA 92037, USA p53 activation by diverse stresses involves post-transla- tional modifications that alter its structure and result in its nuclear accumulation. We will discuss several unresolved topics regarding p53 regulation which are currently under investigation. DNA damage is perhaps the best-studied stress which activates p53, and recent data implicate phosphorylation at N-terminal serine residues as critical in this process. We discuss recent data regarding the potential kinases which modify p53 and the possible role of the resulting phosphorylation events. By contrast, much less is understood about agents which disrupt the mitotic spindle. The cell cycle phase, induction signal, and biochemical mechanism of the reversible arrest induced by microtubule disruption are currently under investigation. Finally, a key event in response to any genotoxic stress is the accumulation of p53 in the nucleus. The factors which determine the steady state level of p53 are starting to be elucidated, but the mechanisms responsible for nuclear accumulation and nuclear export remain controversial. We discuss new studies revealing a mechanism for nuclear retention of p53, and the potential contributions of MDM2 to this process. Keywords: p53; DNA damage; nuclear export signal; microtubule depolymerization; post translational mod- ification Introduction New revelations continue to emerge concerning the mechanisms that control p53 activation in response to a wide range of input signals. This tumor suppressor maintains genetic stability by responding to multiple environmental stresses including DNA damage, ribo- nucleotide depletion, microtubule disruption, redox modulation, cell adhesion, and hypoxia, as well as ‘genetic’ stresses created by activated oncogenes (reviewed in Giaccia and Kastan, 1998). These diverse stimuli appear to invoke a similar set of responses to achieve p53 activation: p53 must first accumulate in the nucleus, and then bind to DNA as a tetramer to transcriptionally regulate a growing list of target genes including p21, GADD45, cyclin G, 14-3-3sigma, thrombospondin 1, MDM2, IGF-BP3, and bax (reviewed in El-Deiry, 1998; Gottlieb and Oren, 1996; Ko and Prives, 1996), as well as c-fos (Elkeles et al., 1999) and others. However, depending on the stress and cell type, p53 activation can lead to responses as dierent as a reversible cycle arrest, induction of a senescent-like state, or apoptosis. Clearly, the induction of the irreversible states of senescence or apoptosis requires a permanent activation of p53 which commits the cell to exit the cycle forever. On the other hand, some agents, such as nocodazole and colcemid, cause a reversible arrest. This raises the important question of whether stimuli that induce cell cycle exit induce dierent alterations in p53 than those that result in a reversible response, which could result in regulation of dierent downstream genes. We infer that there must also be attenuation mechanisms that reverse p53 function subsequent to removal of certain stresses. Since p53 has the power to either target a cell for death or allow it to survive, it must be under rigorous and complex control. p53 contains an N-terminal transactivation domain, a core DNA binding domain, and a C-terminal multifunctional regulatory domain (reviewed in Ko and Prives, 1996). Highly conserved residues in its N- and C-terminal domains are targets for potential post-translational modification via phos- phorylation, dephosphorylation or acetylation (re- viewed in Giaccia and Kastan, 1998). We will discuss how post-translational modification may influence p53 function in a variety of ways. The possibilities include eects on binding to other proteins, such as co- activators or negative regulators, and induction of higher order structural changes mediated by highly conserved amino acid motifs encoded within the primary sequence. For example the multifunctional C-terminus contains a newly identified nuclear export signal (NES) within the tetramerization domain (Stommel et al., 1999). Data are discussed which indicate that the NES is occluded in the p53 tetramer, the active DNA binding conformation. N-terminal phosphorylation and p53 activation p53 protein is translocated to the nucleus, stabilized and rendered competent for DNA binding and transactivation in response to DNA damage. Many studies have attempted to correlate the contributions of p53 protein accumulation, post-translational modifica- tion, and binding to regulatory proteins with the ability of p53 to bind DNA and transactivate downstream target genes. p53 transactivation appears to require its interaction with components of TFIID, including the TATA box binding protein (TBP) and TBP-associated factors (TAFs), via the p53 N-terminal transactivation domain (Lu and Levine, 1995; Thut et al., 1995; Xiao et al., 1994). More recently, transcriptional co- activators p300 and CREB-binding protein (CBP) have also been shown to enhance p53 transactivation *Correspondence: GM Wahl Oncogene (1999) 18, 7656 – 7665 ª 1999 Stockton Press All rights reserved 0950 – 9232/99 $15.00 http://www.stockton-press.co.uk/onc