The number of different mushroom species on earth is estimated at 140,000 of which maybe only 10% are known. Meanwhile, of those ~14,000 species currently recorded, ~50% are considered to possess varying degrees of edibility (17
). Oyster mushrooms (Pleurotus
species) are edible mushrooms and are widespread throughout the hardwood forests of the world (18
). Pleurotus ostreatus
is an edible mushroom widely researched for a variety of properties including antitumor effect. However, the molecular mechanism of its action on colon and breast cancer is not fully understood. Here, we demonstrated that P. ostreatus
induces G0/G1 cell cycle arrest of MCF-7 cells by the up-regulation of the expression of p53 and p21, whereas cell cycle arrest of HT-29 is induced at G0/G1 by the up-regulation of the expression of p21.
Deregulated cell cycle progression is a common abnormality observed in human cancers. Progression through the cell-division cycle is regulated by the coordinated activities of cyclin/cyclin-dependent kinases (CDK) complexes. One level of regulation of these cyclin-CDK complexes is provided by their binding to CDK inhibitors (CKIs)(19
). The CDK inhibitor p21 binds and inhibits the cyclin D-, E-, and A-dependent kinases, regulating G1
to S-phase transition of the cell cycle (20
). Overexpression of p21 is reported to inhibit the proliferation of mammalian cells (21
). In the present study, we observed by using cDNA microarray technology and Western blot analysis that P. ostreatus
up-regulated expression of p21 and p53 in MCF-7 cells which have wild-type p53. Thus, treatment of MCF-7 cells by P. ostreatus
may lead to the induction of p21 by p53 dependent mechanisms in MCF-7 cells. The increase of p53 protein levels in MCF-7 cells treated with P. ostreatus
suggests that its growth inhibitory properties may involve the induction of DNA damage response. Moreover, we have found up-regulation of p21 in HT-29 cells at transcription levels and as well as on a protein level. As previously demonstrated, transcription of the p21 gene may be regulated by p53-dependent as well as p53-independent mechanisms (23
). Our data indicated that P. ostreatus
induces p21 expression in a dose-dependent manner in HT-29 cells, which lack functional p53, suggesting an independent mechanism. The p53 gene product is known to be a key player in the genotoxic-stress response in mammalian cells by inducing the transcription of p21, which in turn inhibits the cyclin E/CDK2-dependent phosphorylation of retinoblastoma (pRb)(24
). Phosphorylation of Rb proteins disrupts binding to the E2F proteins, which allows them to act as transcription activators. E2F targets genes that are up-regulated as a result of Rb phosphorylation encode genes necessary for the completion of the G1/S phase transition, as well as genes necessary for DNA replication (25
). In the present study, P. ostreatus
dose dependently decreased the pRb level, indicating that up-regulation of p21 by P. ostreatus
leads to inhibition of CDK activity, which results in decreased phosphorylation of CDK substrates. These results indicate that the decreased pRb contributes to the G1/S arrest observed in P. ostreatus
treated cells. Moreover, cDNA microarray data showed decreased expression of E2F-1 and TFDP-1, members of the E2F transcription factor family in P. ostreatus
treated MCF-7 and HT-29 cells, suggesting that the decrease in E2F-1 and TFDP-1 levels may also contribute to cell cycle arrest at the G0/G1 phase. In addition, p21 may also inhibit cell cycle progression via the interaction with proliferating cell nuclear antigen (PCNA) (26
). PCNA protein is one of the molecules central in determining the life and death of the cell (28
). The C terminus of p21, containing its nuclear localization signal, binds to and inhibits PCNA, thereby blocking DNA replication (29
). The present study demonstrated that P. ostreatus
slightly decreased the protein level of PCNA in MCF-7 treated cells, which may also contribute to the decreased DNA synthesis. However, expression of PCNA in HT-29 treated cells was not affected by P. ostreatus
In conclusion, our data suggest that P. ostreatus significantly inhibits proliferation of human breast cancer cells MCF-7 and colon cancer cells HT-29 by G0/G1 cell cycle arrest. Therefore, P. ostreatus induces cell cycle arrest of MCF-7 by the up-regulation of the expression of p53 and p21, whereas cell cycle arrest of HT-29 is induced by the up-regulation of the expression of p21 ().
Schematic representation of the expression of cell cycle regulatory genes modulated by P. ostreatus in MCF-7 and HT-29 cells.
In summary, our data demonstrate that the dietary mushroom P. ostreatus specifically inhibits growth of colon and breast cancer cells without significant effect on normal cells, and has a potential therapeutic/preventive effect on breast and colon cancer.