Fidelity of Target Site Duplication and Sequence Preference during Integration of Xenotropic Murine Leukemia Virus-Related Virus

Abstract

Xenotropic murine leukemia virus (MLV)-related virus (XMRV) is a new human retrovirus associated with prostate cancer and chronic fatigue syndrome. The causal relationship of XMRV infection to human disease and the mechanism of pathogenicity have not been established. During retrovirus replication, integration of the cDNA copy of the viral RNA genome into the host cell chromosome is an essential step and involves coordinated joining of the two ends of the linear viral DNA into staggered sites on target DNA. Correct integration produces proviruses that are flanked by a short direct repeat, which varies from 4 to 6 bp among the retroviruses but is invariant for each particular retrovirus. Uncoordinated joining of the two viral DNA ends into target DNA can cause insertions, deletions, or other genomic alterations at the integration site. To determine the fidelity of XMRV integration, cells infected with XMRV were clonally expanded and DNA sequences at the viral-host DNA junctions were determined and analyzed. We found that a majority of the provirus ends were correctly processed and flanked by a 4-bp direct repeat of host DNA. A weak consensus sequence was also detected at the XMRV integration sites. We conclude that integration of XMRV DNA involves a coordinated joining of two viral DNA ends that are spaced 4 bp apart on the target DNA and proceeds with high fidelity.

Analysis of Single-Nucleotide Polymorphisms in Patient-Derived Retrovirus Integration Sites Reveals Contamination from Cell Lines Acutely Infected by Xenotropic Murine Leukemia Virus-Related Virus

“…The SNP rs72789205 allele contains a T in the integration site and heterozygous (C/T) in DU145 and 22Rv1 DNA, compared to C in the patient. EU981808 is identical to an integration site, GU816103, found in an XMRV-infected DU145 clonal cell line (3,6). Therefore, our SNP results provide direct evidence that this integration site is the result of contamination from infected DU145 cells.…”

“…b EU981808 and EU981810 are identical to integration sites GU816103 and EU981678, respectively, from experimentally infected DU145 cells (3,5,6).…”

“…Integration of proviral DNA into the host cell genome is a defining feature of retroviral replication. We previously used a linker-mediated PCR method (5) to sequence and map a total of 14 unique provirus integration sites in human prostate DNA from nine different prostate cancer patients (5,6), supporting XMRV as a bona fide human retrovirus. A recent BLAST analysis showed that two of the patient-derived integration sites are identical to integration sites found in experimental infections of the prostate cancer cell line DU145 (3).…”

“…While the majority of integration sites lack SNPs and are therefore noninformative, our analyses indicate that the proviruses designated EU981808 (VP363) and EU981801 (VP432) originated not from the patient samples but from XMRV-infected cell lines. At the time of the original study on XMRV integration sites (1,5,6), the UCLA laboratory where the integration site mapping was performed did not culture prostate cancer cell lines nor handle mouse strains, making it unlikely that XMRV or XMRV-like sequences would have been present. However, in addition to the work with the human prostate samples, we performed analyses of XMRV integration sites in genomic DNA isolated from experimentally infected DU145 cells and clones (5,6).…”

Is a Retrovirus (XMRV) Risk Factor for Prostate Cancer in Indian Population

“…We need to understand its integration site in host genome, transmission routes, and its interaction with host immune system. Previously, various molecular approaches were focused on the detection of XMRV infection and integration site in the prostate carcinogenesis (Arnold et al, 2010;Kim et al, 2010) but still it is not clear yet.…”