An unconventional role of BMP-Smad1 signaling in DNA damage response: a mechanism for tumor suppression.

The genome is under constant attack by self-produced reactive oxygen species and genotoxic reagents in the environment. Cells have evolved a DNA damage response (DDR) system to sense DNA damage, to halt cell cycle progression and repair the lesions, or to induce apoptosis if encountering irreparable damage. The best studied DDR pathways are the PIKK-p53 and PIKK-Chk1/2. Mutations in these genes encoding DDR molecules usually lead to genome instability and tumorigenesis. It is worth noting that there exist unconventional pathways that facilitate the canonical pathways or take over in the absence of the canonical pathways in DDR. This review will summarize on several unconventional pathways that participate in DDR with an emphasis on the BMP-Smad1 pathway, a known regulator of mouse development and bone remodeling.

Smad1 stabilization and delocalization in response to the blockade of BMP activity.

Signaling at the plasma membrane receptors is generally terminated by some form of feedback regulation, such as endocytosis and/or degradation of the receptors. BMP-Smad1 signaling can also be attenuated by BMP-induced expression of the inhibitory Smads, which are negative regulators of Smad1 transactivation activity and/or BMP antagonists. Here, we report on a novel Smad1 regulation mechanism that occurs in response to the blockade of BMP activity. Lowering the serum levels or antagonizing BMPs with noggin led to upregulation of Smad1 at the protein level in several cell lines, but not to upregulation of Smad5, Smad8 or Smad2/3. The Smad1 upregulation occurs at the level of protein stabilization. Upregulated Smad1 was relocalized to the perinuclear region. These alterations seem to affect the dynamics and amplitude of BMP2-induced Smad1 reactivation. Our findings indicate that depleting or antagonizing BMPs leads to Smad1 stabilization and relocalization, thus revealing an unexpected regulatory mechanism for BMP-Smad1 signaling.

A crucial role for bone morphogenetic protein-Smad1 signalling in the DNA damage response.

DNA damage and the elicited cellular response underlie the etiology of tumorigenesis and ageing. Yet, how this response integrates inputs from cells' environmental cues remains underexplored. Here we report that the BMP-Smad1 pathway, which is essential for embryonic development and tissue homeostasis, has an important role in the DNA damage response and oncogenesis. On genotoxic stress, Atm phosphorylates BMPs-activated Smad1 in the nucleus on S239, which disrupts Smad1 interaction with protein phosphatase PPM1A, leading to enhanced activation and upregulation of Smad1. Smad1 then interacts with p53 and inhibits Mdm2-mediated p53 ubiquitination and degradation to regulate cell proliferation and survival. Enhanced Smad1 S239 phosphorylation, and Smad1 mutations causing S239 substitution were detected in oesophageal and gastric cancer samples, respectively. These findings suggest that BMP-Smad1 signalling participates in the DNA damage response via the Atm-p53 pathway, thus providing a molecular mechanism whereby BMP-Smad1 loss-of-function leads to tumorigenesis, for example, juvenile polyposis and Cowden syndromes.

S6K1 is a multifaceted regulator of Mdm2 that connects nutrient status and DNA damage response.

p53 mediates DNA damage-induced cell-cycle arrest, apoptosis, or senescence, and it is controlled by Mdm2, which mainly ubiquitinates p53 in the nucleus and promotes p53 nuclear export and degradation. By searching for the kinases responsible for Mdm2 S163 phosphorylation under genotoxic stress, we identified S6K1 as a multifaceted regulator of Mdm2. DNA damage activates mTOR-S6K1 through p38alpha MAPK. The activated S6K1 forms a tighter complex with Mdm2, inhibits Mdm2-mediated p53 ubiquitination, and promotes p53 induction, in addition to phosphorylating Mdm2 on S163. Deactivation of mTOR-S6K1 signalling leads to Mdm2 nuclear translocation, which is facilitated by S163 phosphorylation, a reduction in p53 induction, and an alteration in p53-dependent cell death. These findings thus establish mTOR-S6K1 as a novel regulator of p53 in DNA damage response and likely in tumorigenesis. S6K1-Mdm2 interaction presents a route for cells to incorporate the metabolic/energy cues into DNA damage response and links the aging-controlling Mdm2-p53 and mTOR-S6K pathways.

p53 Deficiency leads to compensatory up-regulation of p16INK4a.

p53-p21-cyclin-dependent kinase and p16(INK4a)-cyclin-dependent kinase pathways have parallel functions in preventing tumorigenesis. In cancer patients, tumor suppressor p53 is frequently inactivated through mutations, whereas p16(INK4a) is silenced through promoter methylation. However, the interaction between these two pathways is less well understood. Here, we report that p53 controls p16(INK4a) expression in a unique way. p53 deficiency led to up-regulation of p16(INK4a) in primary mouse embryonic fibroblasts, osteoblasts, and various mouse organs, and an increase in the p16(INK4a) promoter activity, without affecting the half-life of p16(INK4a). Reconstitution of p53, but not mutant p53, restored the proper expression of p16(INK4a). These results indicate that p53 is necessary in repressing p16(INK4a) expression. However, up-regulation of p53 in response to genotoxic stress or nutlin-3 treatment did not down-regulate p16(INK4a). p53 did not repress the p16(INK4a) promoter activity either. These findings suggest that p53 has a necessary but not sufficient role in repressing p16(INK4a) expression. p16(INK4a) elevation in p53(-/-) cells is, at least partially, mediated by Ets1, a known positive regulator of p16(INK4a), as p53 deficiency up-regulated Ets1 through protein stabilization and knockdown of Ets1 down-regulated p16(INK4a) expression in p53(-/-) mouse embryonic fibroblasts. These studies uncover a compensatory mechanism for the loss of p53 and provide a basis for targeting both p53 and p16(INK4a) in cancer therapy.

Ribosomal protein S27-like, a p53-inducible modulator of cell fate in response to genotoxic stress.

Activation of the p53 tumor suppressor upon DNA damage elicits either cell cycle arrest or apoptosis, and the precise mechanism governing cell fate after p53 response has not been well defined. Through genomic analysis, we have identified the ribosomal protein S27-like (RPS27L) as a novel p53 transcriptional target gene. Although RPS27L mRNA levels were consistently induced after diverse p53 activating signals, its change in protein level was stimuli-dependent: it was up-regulated when cells were arrested in response to DNA-damaging agents Adriamycin or VP16 but was down-regulated when cells underwent apoptosis in response to antimetabolite agent 5-fluorouracil. RPS27L is a nuclear protein that forms nuclear foci upon DNA damage. Depletion of RPS27L resulted in deficiency in DNA damage checkpoints, leading to conversion of DNA damage-induced p53 response from cell cycle arrest to apoptosis. We further show that RPS27L positively regulates p21 protein expression. Through this mechanism, RPS27L induction by p53 facilitates p21-mediated cell cycle arrest and protects against DNA damage-induced apoptosis. Thus, RPS27L modulates DNA damage response and functions as a part of the control switch to determine cell fate to DNA damage-p53 response.

Partial sleep deprivation compromises gastric mucosal integrity in rats.

The gastric mucosa is most susceptible to stress that has been shown to induce mucosal damage in humans and animals. This study aims to explore the underlying mechanisms of partial sleep deprivation, as a source of psychophysiological stress, on gastric functions and its effect on mucosal integrity. Sprague-Dawley rats were partially sleep deprived (PSD) for 7 or 14 days by housing inside slowly rotating drums. Gastric tissues and plasma were sampled at the end of the sleep deprivation periods and mucosal lesion scores were evaluated. Morphological examination was performed after Hematoxylin and Eosin staining. Plasma levels of noradrenaline, adrenaline, gastrin, histamine and somatostatin were determined with enzyme immunoassays. Gastric acidity was measured with acid-base titration in pylorus ligated rats. Gastric mucosal blood flow was evaluated with Laser Doppler Flowmetry. It was found that gastric lesions were induced in about 30%-50% of the PSD rats. Gastric acidity as well as plasma levels of noradrenaline, gastrin and histamine were elevated. Gastric mucosal blood flow and plasma somatostatin level were on the contrary reduced, especially in rats with PSD for 14 days. It is concluded that partial sleep deprivation compromises gastric mucosal integrity by increasing gastric acidity, plasma levels of noradrenaline, gastrin, histamine, and decreasing gastric mucosal blood flow. These results provided experimental evidence on the gastric damaging effects of PSD and it could be one of the risk factors contributing to gastric ulcer formation.