Chronic p53-independent p21 expression causes genomic instability by deregulating replication licensing

Abstract

The cyclin-dependent kinase inhibitor p21(WAF1/CIP1) (p21) is a cell-cycle checkpoint effector and inducer of senescence, regulated by p53. Yet, evidence suggests that p21 could also be oncogenic, through a mechanism that has so far remained obscure. We report that a subset of atypical cancerous cells strongly expressing p21 showed proliferation features. This occurred predominantly in p53-mutant human cancers, suggesting p53-independent upregulation of p21 selectively in more aggressive tumour cells. Multifaceted phenotypic and genomic analyses of p21-inducible, p53-null, cancerous and near-normal cellular models showed that after an initial senescence-like phase, a subpopulation of p21-expressing proliferating cells emerged, featuring increased genomic instability, aggressiveness and chemoresistance. Mechanistically, sustained p21 accumulation inhibited mainly the CRL4-CDT2 ubiquitin ligase, leading to deregulated origin licensing and replication stress. Collectively, our data reveal the tumour-promoting ability of p21 through deregulation of DNA replication licensing machinery-an unorthodox role to be considered in cancer treatment, since p21 responds to various stimuli including some chemotherapy drugs.

p21WAF1/Cip1 - PCNA: Fatal attraction

“…Indeed analyzing the data in our report, we found out that this was the case. 5 TLS is the main DNA damage tolerance pathway of the cell representing an important "defense" line, as it mends the lesions that escape from the surveillance of the error-free DNA repair mechanisms, allowing completion of chromosome replication. 7 Apparently, prolonged p21 WAF1/Cip1 expression could lead to reduced TLS activity contributing together with re-replication to replication fork stalling, collapse and generation of double strand breaks fuelling genomic instability (Fig.…”

“…In line with this notion, we showed that protracted p21 WAF1/Cip1 expression abrogated the property of PCNA to regulate the degradation cycle of the replication licensing factors Cdt1 and Cdc6, 6 triggering re-replication. 5 Consecutively, p21…”

“…4 Within this context we demonstrated that the duration of p21 WAF1/Cip1 expression becomes longer (chronic), as opposed to the short-term p53-mediated induction (transient cell cycle arrest), leading to an initial senescent phase that a first glance appears as an efficient anti-proliferative and eventually anti-tumor response. 5 Although cellular senescence is considered an anti-tumor barrier, in reality it is an "imperfect" tumor-suppressive mechanism. The prevailing view for the later notion is the so called senescence-associated secretory phenotype (SASP), a process that exercises pro-tumorigenic effects.…”

“…The selective process is actually an extensive error-prone repair process that ensures that over time the "fittest and more stable cells" will survive and emerge. 5 An essential biochemical act assisting in p21 WAF1/Cip1 senescence bypass is suppression of the INK4/ARF locus by the deregulated replication licensing factor Cdc6, 6 rendering the senescent phase reversible. 2 Another consequence of continuous p21 WAF1/Cip1 expression, related to PCNA-dependent functions, could be suppression of PCNA mono-ubiquitinilation (data not shown), essential for translesion DNA synthesis (TLS) and repair.…”

“…2 We found out, however, that p21 WAF1/Cip1 -driven senescence conceals a more complex and "malicious" program characterized by S-M phase dissociation, due to mitotic blockage and over-replication in the form of re-replication, as a result of deregulated licensing. 5 The key biochemical event leading to aberrant licensing is the interaction of p21 WAF1/Cip1 with Proliferating Cell Nuclear Antigen (PCNA), a DNA sliding clamp involved in fundamental cell cycle functions such as, DNA replication and repair. Among all PCNA-interacting partners, p21 WAF1/Cip1 possesses the highest binding affinity (K D »2.5 nM), implying that under conditions of "chronic" expression, p21…”