Wang et al. Journal of Experimental & Clinical Cancer Research 2010, 29:140 http://www.jeccr.com/content/29/1/140 Page 6 of 7 primary breast cancer (n = 112) by IHC analysis, and showed that HBO1 was highly expressed in breast can- cer (Table 1) and positively correlated with ERa (p < 0.001) and PR (p = 0.002). Moreover, HBO1 protein level correlated positively with histology grade in ER positive tumors (p = 0.016) rather than ERa negative tumors through statistical analysis. As a coactivator of the replication licensing factor Cdt1 [17], HBO1 belongs to one component of the Replication Initiation Proteins known as prereplicative complex (pre-RC) proteins. Sev- eral pre-RC proteins are over-expressed in cancer and serve as good tumor markers. And some of them, such as Cdc6 and Cdt1, are elevated by E2 treatment in breast cancer. To determine whether HBO1 was also affected by E2, quantitative real-time PCR and western blot were performed. The results suggested HBO1 was elevated after E2 treatment. Further study demonstrated the E2-induced HBO1 upregulation could be inhibited by ICI 182,780 as well as ERa siRNA. Estrogen receptor (ER) signaling is involved in many human diseases such as breast cancer [18], lung cancer [19,20] and cardiovascular diseases [21,22]. It was reported that E2 activated the signal-transducing ERK1/2 pathway, which were critical for cell proliferation [23-25], in human mammary cancer-derived cell lines, MCF-7 and T47 D [26-28]. We explored whether ERK1/2 signal- ing pathway was involved in the expression of HBO1 increased by E2. The MEK1/2 inhibitor U0126 signifi- cantly inhibited the expression of HBO1 in T47 D and MCF-7 cells, suggesting that E2 increased the expression of HBO1 through the ERK1/2 signaling pathway. Since previous studies have shown that progesterone receptor activates the Src/p21ras/ERK pathway via cross-talk with estrogen receptor in breast cancer [29], the positive cor- relation between HBO1 protein levels and PR which we obtained in statistical analysis was reasonable. Estrogen exposure has been regarded as high risk factor of breast cancer. It has been reported that HBO1 strongly enhanced ER-mediated transcription [9], which indicated that HBO1 might play a role in the progress of breast cancer. In this study, we proved E2 could upregulate the mRNA and protein level of HBO1. Meanwhile, knock- down of ERa with siRNA significantly inhibited the upre- gulation of HBO1, indicating that cross-talking was happening between ERa and HBO1 in breast cancer, the biological functions of which need to be further studied. Conclusion Our data have demonstrated that the HBO1 protein levels correlated positively with ERa (p < 0.001) and PR (p = 0.002) expression in breast cancer. Further more, HBO1 protein levels correlated positively with histology grade in ERa positive tumors (p = 0.016). We also showed that the ERK1/2 signaling pathway was involved in the expression of HBO1 increased by E2. These find- ings suggested a potential for targeting HBO1 as a novel means of breast cancer therapy as well as a potential diagnosis marker for ERa positive breast cancer. Acknowledgements This work is supported by grants from National Natural Science Funds: 30770426, 30930025, 30900266 and 31000348; State Key Project Specialized for Infectious Diseases: 2008ZX10002-015, 2008ZX10002-021, 2008ZX10001-02 and 2008ZX10004-014; National Basic Research Program of China (973 Program): 2010CB912100 (2010CB912104); National Fundamental Fund Project: J0730860; Shanghai Leading Academic Discipline Project: B110. Author details 1Department of Biochemistry and Molecular Biology, Shanghai Medical College of Fudan University, Shanghai, 200032, China. 2Institutes of Biomedical Science of Fudan University, Shanghai, 200032, China. 3Institute of Nautical Medicine, Nantong University, Nantong, 226001, China. Authors’ contributions WWZ carried out the design of the study, performed IHC, real-time PCR, drafted the manuscript. LHO performed the western blot. WYH participated in SPSS Statistical Analysis. LYH participated in IHC and IOD scoring. WWB participated in real-time PCR and cell culture. ZL performed SPSS Statistical Analysis. HY and YJW participated in IHC. SLL participated in collection of breast cancer specimens. XJJ participated in the design of the study, drafted the figure. YXJ performed the design of the study, and helped drafting the manuscript. GJX performed the collection of breast cancer specimens. All authors read and approved the final manuscript. 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