Background
Polycomb group (PcG) proteins are evolutionary conserved gene silencers, which play an important role in the development of vertebrate organisms. These proteins regulate cell proliferation, senescence and tumorigenesis via well-known growth regulatory pathways [
1]. Overexpression of key PcG proteins such as BMI1 has been found in several human malignancies including breast cancer, colorectal cancer, nasopharyngeal carcinoma, melanoma, gastric cancer, and bladder cancer [
2‐
7]. Overexpression of BMI1 often correlates with poorer prognosis [
4,
6,
7]. BMI1 also plays an important role in self-renewal of hematopoietic stem cells (HSCs), neural stem cells and mammary stem cells [
8‐
10].
After a finite number of cell divisions, most normal human cells undergo cellular senescence, which constitutes a powerful barrier to oncogenesis[
11]. Overexpression of BMI1 has been shown to bypass this barrier in human mammary epithelial cells and fibroblasts [
12,
13]. In addition to BMI1, mammalian cells also express a BMI1-related PcG protein Mel-18 (also known as polycomb group ring finger 2 (PCGF2)). The Mel-18 gene product is structurally highly similar to BMI1 protein. Interestingly, we have found that BMI1 is negatively regulated by Mel-18 and that Mel-18 is overexpressed in senescent fibroblasts. Accordingly, Mel-18 overexpression leads to accelerated or premature senescence in proliferating fibroblasts by repression of BMI1 [
14]. Similar to human fibroblasts, expression of Mel-18 negatively correlates with BMI1 in a number of breast cancer cell lines and breast tumors [
15]. Negative correlation between BMI1 and Mel-18 expression was also recently reported in hematopoietic stem cells [
16].
We also reported that Mel-18 overexpression in breast cancer cell line MCF7 results in downregulation of BMI1 and reduction of transformed phenotype. Furthermore, downregulation of BMI1 by Mel-18 overexpression and knockdown of BMI1 expression by RNA interference (RNAi) approach is accompanied by downregulation of AKT/PKB (Protein Kinase B) activity [
15]. Thus our data suggested that Mel-18 acts as a tumor suppressor in breast epithelial cells. Consistent with our data, Lee et al. also recently reported that Mel-18 negatively regulates AKT and that its overexpression inhibits growth of breast cancer cells [
17]. However, the function of Mel-18 is still debatable. It is described as a potential tumor suppressor in some studies [
17‐
19]; while in other studies, it was found that similar to
BMI1,
Mel-18 can act as an oncogene [
20,
21]. The opposite role of BMI1 and Mel-18 in human fibroblasts and breast cancer cells is an interesting finding. However, it needs to be verified in other cell types and pathological conditions. Importantly, the role of Mel-18 in cancers other than breast and prostate cancers is still not clear. Regulation of AKT/PKB pathway by BMI1 and Mel-18 activity also needs further confirmation.
Gastric cancer is one of the most common malignancies throughout the world, and mechanisms that underlie the carcinogenesis of gastric cancer are still poorly understood. It has been reported that BMI1 is overexpressed in gastric cancer cells and is an independent prognosis factor [
6]. However, the exact role of BMI1 in gastric cancer is far from clear. Potential tumor suppressive role of Mel-18 in gastric cancer is also not known. Here, we show that BMI1 is overexpressed in gastric cancer cell lines and gastric tumors, and its expression correlated with advanced clinical stage, lymph node metastasis, and poor prognosis. Importantly, we show that expression of Mel-18 is decreased in gastric cancer and is negatively correlated with the expression of BMI1 in both gastric cancer cells and normal gastric epithelial cells. We also report that downregulation of BMI1 by Mel-18 overexpression and knockdown of BMI1 expression in gastric cell lines is accompanied by downregulation of AKT/PKB activity and upregulation of p16, which resulted in induction of a senescence-like phenotype and reduction of transformed properties in gastric cancer cell lines. These data suggest that
BMI1 acts as an oncogene via regulation of p16 and AKT/PKB, and
Mel-18 acts as a tumor suppressor via downregulation of BMI1 during the development of gastric cancer.
Discussion
Arguably BMI1 is the most well studied PcG protein that is known to regulate proliferation and senescence in mammalian cells. It not only inhibits senescence and immortalizes human mammary epithelial cells (HMECs) [
13], but it can also transform keratinocytes via downregulation of tumor suppressors and differentiation related factors[
25]. It also cooperates with H-Ras to transform HMECs via dysregulation of multiple growth-regulatory pathways [
24], and plays a central role in mediating leukemic transformation and development [
26]. Aberrant expression of BMI1 has been found in several human cancers and its overexpression is often correlated with poor prognosis in many types of cancers[
2‐
7]. Most importantly, BMI1 is required for self-renewal of normal and malignant stem cells [
8‐
10]. Hence, BMI1 is considered an important therapy target [
27,
28]. Because BMI1 is an important regulator of cell proliferation and stem cell phenotype, its own regulators are likely to be very important for onocogenesis and self-renewal of normal and cancer stem cells.
We have previously reported that BMI1 is regulated by another PcG protein Mel-18 in human fibroblasts [
14], and that by doing so Mel-18 can potentially function as a tumor suppressor. Potential tumor suppressor role of Mel-18 has also been suggested in few other studies [
15,
17]. In the present study we provide
in vitro and
in vivo evidences supporting this notion during gastric cancer development. Overexpression of BMI1 in gastric cancer has been previously reported [
6,
29], however the potential mechanism of its overexpression remained unclear. Here we confirmed previous observation that BMI1 is overexpressed in gastric cancer cell lines and gastric tumors and that BMI1 overexpression correlates with poor prognosis.
Importantly, here we show that Mel-18 is downregulated in gastric cancer cells and gastric tumors, and that there is a negative correlation between Mel-18 and BMI1 expression in gastric cancer cells. Thus, our data suggest that overexpression of BMI1 in gastric tumors and cell lines may due to downregulation of Mel-18 in gastric cancer cells. Consistent with its tumor suppressor role, we also found that Mel-18 negatively regulated AKT expression, and induced p16 expression and senescence in gastric cancer cell lines. In a recent study, BMI1 overexpression was closely related with the Lauren's and Borrmann's classification and clinical stage in gastric tumors [
29]. However, in our study we did not find correlation between BMI1 expression and tumor size, T classification or differentiation in gastric tumors, which was not consistent with the
in vitro study that BMI1 regulates proliferation. The discrepancy could be due to the limited number of samples in our study or other interfering factors. However, we did find that BMI1 overexpression positively correlated with lymph node metastasis, and clinical stages of the tumors. Our
in vitro study also showed that overexpression of Mel-18, and knockdown of BMI1 expression, inhibit the ability of migration in gastric cancer cells. It's the first time to find that Mel-18 and Bmi-1 regulate cellular migration in
in vitro model, and provide preliminary direct evidence for the possibility of Mel-18 and Bmi-1 regulate the metastasis of cancer. Collectively, our data suggest that Mel-18 and BMI1 not only play important roles in tumorigenesis, but may also involve in the progression and metastasis of gastric cancer.
It is interesting to note that our data suggest that BMI1 but not the Mel-18 is an independent negative prognosis factor. Patients with high BMI1 expression survived significantly shorter than those with low or no BMI1 expression suggesting that BMI1 is a key regulator and a valuable molecular marker of therapy failure in gastric cancer patients. It also suggest that BMI1 may be regulated by factors other than Mel-18, and that Mel-18 could only partially regulate BMI1, particularly during advanced stages of gastric cancer, which can explain why Mel-18 expression correlated with BMI1 but was not an independent prognosis factor.
Taken together, gastric tumor tissues expressed significantly higher BMI1 and lower Mel-18 compared with normal gastric mucosal tissues, and BMI1 correlated with lymph node metastasis, clinical stages, and prognosis. Hence, detection of Mel-18 and BMI1 expression may be helpful in supporting the diagnosis and determining the prognosis of gastric cancer in clinical practice.
Although
INK4A/ARF locus is the most cancer relevant target of BMI1 and Mel-18, there are several new reports, which suggest that these PcG proteins, in particular BMI1 may regulate tumorigenesis independent of p16 pathway [
17,
24,
25,
30]. Our present data using gastric cancer cells also suggest that BMI1 and Mel-18 can regulate tumorigenesis, cell migration and metastasis at least partially independent of p16, which is a known tumor suppressor involved in regulation of cell proliferation and possibly metastasis [
31‐
33]. Another important target of BMI1 and Mel-18 appears to be AKT [
15], which is known to regulate tumorigenesis and cancer metastasis in several cancers [
34‐
36]. Accordingly, our present study suggests that regulation of gastric tumorigenesis and metastasis by Mel-18 and BMI1 may involve AKT, which was overexpressed in gastric cancer tissues and positively correlated with the depth of invasion and lymph node metastasis in gastric cancer patients.
Competing interests
The authors declare that they have no competing interests.
Authors' contributions
XWZ , YPS, QL, WQ, YWL, YFC perform the experiment. BYL, FCZ, and JL supervised the experiment or provide technique support. XWZ, GPD, and WJG analyzed the data and prepared the manuscript. WJG designed the experiment and supervised the project. All authors read and approved the final manuscript.