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.

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BackgroundSjögren’s syndrome (SS) is a common autoimmune disease in which the main targets of immune injury are specific secretory epithelia, such as salivary and lacrimal glands. Stress appears to play a significant role in the initiation of this entity.ObjectivesThe aim of the present study was to investigate whether chronic stress plays a role in triggering endoplasmic reticulum (ER) stress in salivary gland epithelial cells from SS patients.MethodsMinor salivary gland biopsy specimens were obtained from six SS patients and six control patients with sicca symptoms not fulfilling AECG criteria [1]. The expression and cellular localization of β1-, β2- and α1-adrenoceptors and the levels of cAMP were measured by immunofluorescence. The morphology of the ER was evaluated in situ by Transmission Electron Microscopy (TEM). Primary salivary gland epithelial cell lines (SGEC) derived from minor salivary gland biopsies, were established by the explant out-growth technique [2] and were treated with epinephrine and norepinephrine. The protein levels of the ER stress markers GRP78/Bip and C/EBP homologous protein (CHOP) were determined by immunoblot analysis.Results In situ immunofluorescence staining revealed increased expression of β1-, β2- and α1 adrenoceptors as well as cAMP levels in tissues derived from SS patients compared to controls. TEM evaluation of salivary tissues from SS subjects revealed extensive dilation of the ER lumen compared to controls. Treatment of SGEC with epinephrine and norepinephrine did not influence cell survival (cell viability assay). To mimic chronic stress in vitro, epinephrine was applied for 10 days on SGEC. It was found that 20μM Epinephrine induced severe ER stress on SGEC, as attested by increased expression of GRP78/Bip and CHOP, after 3 and 6 days of treatment. The expression of ER stress markers returned to basal levels after 10 days of treatment.ConclusionOur data revealed that adrenergic receptors are differentially expressed by salivary epithelium of SS patients compared to control tissues. This finding was further substantiated by the increased levels of cAMP, indicating a profound sympathetic stimulation of this secretory tissue. The dilatated ER lumen of the salivary gland epithelial cells from SS patients confirmed the occurrence of ER stress. Moreover, the finding that catecholamines induce severe ER stress in vitro in SGEC under conditions resembling chronic stress suggests a causal relationship between sympathetic tone and ER stress. Deciphering the role of chronic stress in ER machinery will provide important insights that may lead to novel therapeutic targets for SS.References[1] Vitali C, et al. Ann Rheum Dis2002; 61:554–8.[2] Kapsogeorgou EK, et al. J Immunol2001; 166:3107–13.AcknowledgementThe research has been funded in part by the European Union’s Horizon 2020 multicentric protocol HARMONICSS; H2020-SC1-2016; grant agreement no: 731944.Disclosure of InterestsNone declared