Gene Expression and Post-Transcriptional Regulation of DNA Tumor Viruses

Division Of Clinical Sciences - Nci

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Abstract

Human papillomavirus type 16 (HPV16) or 18 (HPV18) infection, acquired primarily via sexual transmission, is widely recognized as a leading cause of cervical, anal, and oropharyngeal cancer. Persistent infection with these oncogenic or high-risk (HR) HPVs leads to HPV DNA integration randomly into the host genome, which plays a critical role in the progression of cervical dysplasia and ultimately the development of cervical cancers. Mouse papillomavirus type 1 (MmuPV1) and cotton-tail rabbit papillomavirus (CRPV) infection models are two gold-standard preclinical animal models for studying HPV-associated infections and tumors. We have constructed a full transcription map of MmuPV1 and CRPV in tumor tissues to guide the field research. Two viral oncogenes, E6 and E7, responsible for HR-HPV carcinogenesis are highly expressed from the integrated viral DNA in the right place in the chromosome. We discovered only one of multiple integrated viral DNA copies expressible for cell transformation and clonal selection in cervical tissues and cervical cancer-derived cell lines for HR-HPV carcinogenesis. Oncogenic HPV E6 and E7 destabilize cellular tumor suppressor proteins and induce aberrant expression of a subset of oncogenic and tumor-suppressive microRNAs (miRNAs) and long noncoding RNAs (lncRNAs) such as lnc-FANCI-2. In HPV16 and HPV18, E6 and E7 are transcribed as a single bicistronic E6E7 RNA bearing 3 exons and 2 introns, with intron 1 in the E6 coding region. Splicing of the intron 1 in E6E7 pre-mRNA disrupts the E6 open reading frame (ORF) but is required to reinitiate translation of the E7 ORF downstream. Thus, RNA splicing regulates the production of viral E6 and E7 and this splicing is controlled by a branch point adenosine at nt 385 in the E6 intron of HPV16 and by two alternative branch point adenosines either at nt 384 or at nt 388 in the E6 intron of HPV18. We found that cellular RNA-splicing factors, including SRSF3 (SRp20) and hnRNP A1, control RNA splicing of the viral early transcripts and viral early-to-late switch through binding to viral RNA cis-elements. We demonstrated that SRp20 is a proto-oncogene essential for cell proliferation by regulating genome-wide RNA splicing of several hundred host genes. When overexpressed, SRp20 induces cell transformation and tumor formation. Together, the HPV infection-induced degradation of host tumor suppressive proteins, aberrant expression of oncogenic and tumor-suppressive miRNAs and lncRNAs, and enhancement of SRp20 expression could be the major cooperative mechanisms in HR-HPV carcinogenesis. Kaposi's sarcoma-associated herpesvirus (KSHV) is a lymphotropic DNA tumor virus that induces Kaposi's sarcoma (KS), primary effusion lymphoma (PEL) or body cavity-based B-cell lymphoma, and multicentric Castleman disease (MCD). Among these malignancies, KS occurs frequently in patients who are infected with HIV. PEL and MCD feature increased levels of cytokines (IL6 and IL10). Latent KSHV infection in KS lesions and PEL-derived B cells can be reactivated as lytic KSHV infection by various stress conditions or inflammation. In KSHV lytic infection, the viral lytic gene ORF57 encodes a multifunctional, caspase-7-sensitive protein to regulate the expression of a subset of viral lytic and host genes at the post-transcriptional level. We solved the ORF57 crystal structure both as a dimer at 3.5 angstroms and as a monomer at 3 angstroms. We demonstrated that ORF57 functions as a dimer in the cells and carries out its functions by binding to RNA and interacting with cellular RNA-binding proteins. As a viral splicing factor first identified in the virology field by our laboratory, ORF57 promotes RNA splicing by interacting with the spliceosomal machinery. ORF57 binds to and stabilizes viral and host RNAs by interacting with host RNA capping, export, and polyadenylation factors to regulate the expressions of both viral and host genes. ORF57 also promotes protein translation by interrupting miRNA-mediated RNA instability and -translational repression. Our recent findings indicate that ORF57 inhibits host innate immunity against lytic KSHV infection by interacting with PACT, PKR, Ago2, and GW182, leading to block formation of stress granules and P-bodies.