Jonathan Low

Gangliosides have been shown to be plasma membrane receptors for both murine polyomavirus and SV40, while JC virus uses serotonin receptors. In contrast, little is known of the membrane receptor and entry pathway for BK virus (BKV), which can cause severe disease in immunosuppressed bone marrow and renal transplant patients. Using sucrose flotation assays, we investigated BKV binding to and interaction with human erythrocyte membranes and determined that this interaction was dependent on a neuraminidase-sensitive, proteinase K-resistant molecule. BKV was found to interact with the gangliosides GT1b and GD1b. The terminal alpha2-8-linked disialic acid motif, present in both of these gangliosides, is likely to be important for this interaction. We also determined that the addition of GD1b and GT1b to LNCaP cells, which are normally resistant to BKV infection, made them susceptible to the virus. In addition, BKV interacted with membranes extracted from the endoplasmic reticulum (ER) and infection was blocked by the addition of brefeldin A, which interferes with transport from the ER to the Golgi apparatus. These data demonstrate that BKV uses the gangliosides GT1b and GD1b as receptors and passes through the ER on the way to the nucleus.

Transcriptional silencing of tumor suppressor genes by DNA methylation plays an important role in tumorigenesis. These aberrant epigenetic modifications may be mediated in part by elevated DNA methyltransferase levels. DNA methyltransferase 1 (DNMT1), in particular, is overexpressed in many tumor types. Recently, we showed that Dnmt1 is transcriptionally regulated by E2F transcription factors and that retinoblastoma protein (pRb) inactivation induces Dnmt1. Based on these observations, we investigated regulation of Dnmt1 by polyomavirus oncogenes, which potently inhibit the pRb pocket protein family. Infection of primary human prostate epithelial cells with BK polyomavirus dramatically induced Dnmt1 transcription following large T antigen (TAg) translation and E2F activation. For in vivo study of Dnmt1 regulation, we used the transgenic adenocarcinoma of the mouse prostate (TRAMP) model, which expresses the SV40 polyomavirus early region, including TAg, under control of a prostate-specific promoter. Analysis of TRAMP prostate lesions revealed greatly elevated Dnmt1 mRNA and protein levels beginning in prostatic intraepithelial neoplasia and continuing through advanced prostate cancer and metastasis. Interestingly, when TRAMP mice were treated in a chemopreventive manner with the DNA methyltransferase inhibitor 5-aza-2'-deoxycytidine (5-aza), 0 of 14 mice developed prostate cancer at 24 weeks of age, whereas 7 of 13 (54%) control-treated mice developed poorly differentiated prostate cancer. Treatment with 5-aza also prevented the development of lymph node metastases and dramatically extended survival compared with control-treated mice. Taken together, these data suggest that Dnmt1 is rapidly activated by pRb pathway inactivation, and that DNA methyltransferase activity is required for malignant transformation and tumorigenesis.